Genomic characterization of cocirculating Corynebacterium diphtheriae and non-diphtheritic Corynebacterium species among forcibly displaced Myanmar nationals, 2017-2019.

Respiratory diphtheria is a serious infection caused by toxigenic Corynebacterium diphtheriae, and disease transmission mainly occurs through respiratory droplets. Between 2017 and 2019, a large diphtheria outbreak among forcibly displaced Myanmar nationals densely settled in Bangladesh was investigated. Here we utilized whole-genome sequencing (WGS) to characterize recovered isolates of C. diphtheriae and two co-circulating non-diphtheritic Corynebacterium (NDC) species - C. pseudodiphtheriticum and C. propinquum. C. diphtheriae isolates recovered from all 53 positive cases in this study were identified as toxigenic biovar mitis, exhibiting intermediate resistance to penicillin, and formed four phylogenetic clusters circulating among multiple refugee camps. Additional sequenced isolates collected from two patients showed co-colonization with non-toxigenic C. diphtheriae biovar gravis, one of which exhibited decreased susceptibility to the first-line antibiotics and harboured a novel 23-kb multidrug resistance plasmid. Results of phylogenetic reconstruction and virulence-related gene contents of the recovered NDC isolates indicated they were likely commensal organisms, though 80.4 %(45/56) were not susceptible to erythromycin, and most showed high minimum inhibition concentrations against azithromycin. These results demonstrate the high resolution with which WGS can aid molecular investigation of diphtheria outbreaks, through the quantification of bacterial genetic relatedness, as well as the detection of virulence factors and antibiotic resistance markers among case isolates.

The Carboxyl Terminal Regions of P0 Protein Are Required for Systemic Infections of Poleroviruses.

P0 proteins encoded by poleroviruses Brassica yellows virus (BrYV) and Potato leafroll virus (PLRV) are viral suppressors of RNA silencing (VSR) involved in abolishing host RNA silencing to assist viral infection. However, other roles that P0 proteins play in virus infection remain unclear. Here, we found that C-terminal truncation of P0 resulted in compromised systemic infection of BrYV and PLRV. C-terminal truncation affected systemic but not local VSR activities of P0 proteins, but neither transient nor ectopic stably expressed VSR proteins could rescue the systemic infection of BrYV and PLRV mutants. Moreover, BrYV mutant failed to establish systemic infection in DCL2/4 RNAi or RDR6 RNAi plants, indicating that systemic infection might be independent of the VSR activity of P0. Partially rescued infection of BrYV mutant by the co-infected PLRV implied the functional conservation of P0 proteins within genus. However, although C-terminal truncation mutant of BrYV P0 showed weaker interaction with its movement protein (MP) when compared to wild-type P0, wild-type and mutant PLRV P0 showed similar interaction with its MP. In sum, our findings revealed the role of P0 in virus systemic infection and the requirement of P0 carboxyl terminal region for the infection.

Molecular Detection of Potato Viruses in Bangladesh and Their Phylogenetic Analysis.

Potato (Solanum tuberosum) is a major food source in the whole world including Bangladesh. Viral diseases are the key constraint for sustainable potato production by reducing both quality and quantity. To determine the present status of eight important potato viruses in Bangladesh, tuber samples were collected from three major potato growing regions (Munshiganj, Jessore and Bogra districts) in January-February 2017 and February 2018. Reverse transcription polymerase chain reaction (RT-PCR) with coat protein (CP)-specific primers were used to amplify CP sequences of the respective viruses, and confirmed by sequencing, which were deposited in the GenBank. Results indicated that the tuber samples were subjected to Potato leafroll virus (PLRV), Potato virus X (PVX), Potato virus Y (PVY), Potato virus S (PVS), Potato virus H (PVH), Potato aucuba mosaic virus (PAMV) and Potato virus M (PVM) infection, whereas mixed infections were very common. Phylogenetic analysis revealed that the PLRV from this study was closely related to a Canadian and a Chinese isolate, respectively; PVX was closely related to a Canadian and a Chinese isolate, respectively; PVY was closely related to a Chinese isolate; PVS was closely related to a Chinese and an Iranian isolate, respectively; PAMV was closely related to a Canadian isolate; PVH was closely related to a Huhhot isolate of China; and PVM was closely related to an Indian and an Iranian isolate, respectively. As far as we know, PAMV in this study is the first report in Bangladesh. These findings will provide a great scope for appropriate virus control strategies to virus free potato production in Bangladesh.

The Three Essential Motifs in P0 for Suppression of RNA Silencing Activity of Potato leafroll virus Are Required for Virus Systemic Infection.

Higher plants exploit posttranscriptional gene silencing as a defense mechanism against virus infection by the RNA degradation system. Plant RNA viruses suppress posttranscriptional gene silencing using their encoded proteins. Three important motifs (F-box-like motif, G139/W140/G141-like motif, and C-terminal conserved region) in P0 of Potato leafroll virus (PLRV) were reported to be essential for suppression of RNA silencing activity. In this study, Agrobacterium-mediated transient experiments were carried out to screen the available amino acid substitutions in the F-box-like motif and G139/W140/G141-like motif that abolished the RNA silencing suppression activity of P0, without disturbing the P1 amino acid sequence. Subsequently, four P0 defective mutants derived from a full-length cDNA clone of PLRV (L76F and W87R substitutions in the F-box-like motif, G139RRR substitution in the G139/W140/G141-like motif, and F220R substitution in the C-terminal conserved region) were successfully generated by reverse PCR and used to investigate the impact of these substitutions on PLRV infectivity. The RT-PCR and western blot analysis revealed that these defective mutants affected virus accumulation in inoculated leaves and systemic movement in Nicotiana benthamiana as well as in its natural hosts, potato and black nightshade. These results further demonstrate that the RNA silencing suppressor of PLRV is required for PLRV accumulation and systemic infection.

Post-Kala-Azar Dermal Leishmaniasis Without Previous History of Visceral Leishmaniasis.

Post-kala-azar dermal leishmaniasis (PKDL) without previous visceral leishmaniasis (VL) is a rare dermatological manifestation of Leishmania infection. To date, most of the reported cases neither showed parasitological confirmation nor explained the outcome of treatment. Herein, we report three confirmed cases that were were successfully cured after miltefosine treatment.