Comparative genome characterization of Leptospira interrogans from mild and severe leptospirosis patients

Leptospirosis is a zoonotic disease caused by spirochetes from the genus Leptospira. In Thailand, Leptospira interrogans is a major cause of leptospirosis. Leptospirosis patients present with a wide range of clinical manifestations from asymptomatic, mild infections to severe illness involving organ failure. For better understanding the difference between Leptospira isolates causing mild and severe leptospirosis, illumina sequencing was used to sequence genomic DNA in both serotypes. DNA of Leptospira isolated from two patients, one with mild and another with severe symptoms, were included in this study. The paired-end reads were removed adapters and trimmed with Q30 score using Trimmomatic. Trimmed reads were constructed to contigs and scaffolds using SPAdes. Cross-contamination of scaffolds was evaluated by ContEst16s. Prokka tool for bacterial annotation was used to annotate sequences from both Leptospira isolates. Predicted amino acid sequences from Prokka were searched in EggNOG and David gene ontology database to characterize gene ontology. In addition, Leptospira from mild and severe patients, that passed the criteria e-value < 10e-5 from blastP against virulence factor database, were used to analyze with Venn diagram. From this study, we found 13 and 12 genes that were unique in the isolates from mild and severe patients, respectively. The 12 genes in the severe isolate might be virulence factor genes that affect disease severity. However, these genes should be validated in further study.

SARS-CoV-2 RNA shedding in recovered COVID-19 cases and the presence of antibodies against SARS-CoV-2 in recovered COVID-19 cases and close contacts

Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). COVID-19 emerged in December 2019 and has spread globally. Although Thailand has been effective at controlling the spread of COVID-19, disease surveillance and information on antibody responses in infected cases and close contacts are needed because there is still no specific treatment or vaccine available. We investigated 217 recovered COVID-19 cases to monitor their viral RNA shedding and production of antibodies against SARS-CoV-2. The presence of antibodies in blood samples from 308 close contacts of COVID-19 cases was also determined. Viral RNA was still detectable in 6.6 % of recovered COVID-19 cases. The most prolonged duration of viral RNA shedding detected in this study was 105 days. IgM, IgG, and IgA antibodies against SARS-CoV-2 were detected in 13.82, 88.48, and 83.41 % of the recovered cases 4-12 weeks after disease onset, respectively. Although the patients had recovered from their illness, the levels of antibodies detected showed association with their symptoms during their stay in hospital. Fifteen of the 308 contacts (4.87 %) of COVID-19 cases tested positive for IgG antibodies. The presence of antibodies against SARS-CoV-2 suggested that there was viral exposure among close contacts. Viral clearance and the pattern of antibody responses in infected individuals are both crucial for effectively combatting SARS-CoV-2. Our study provides additional information on the natural history of this newly emerging disease related to both natural host defenses and a strategy for vaccine development.

Endotoxemia and circulating bacteriome in severe COVID-19 patients

PurposeWhen severe, COVID-19 shares many clinical features with bacterial sepsis. Yet, secondary bacterial infection is uncommon. However, as epithelium are injured and barrier function is lost, bacterial products entering the circulation might contribute to the pathophysiology of COVID-19. MethodsWe studied 19 adults, severely ill patients with COVID-19 infection, who were admitted to King Chulalongkorn Memorial Hospital, Bangkok, Thailand, between 13th March and 17th April 2020. Blood samples on day 1, 3, and 7 of enrollment were analyzed for endotoxin activity assay (EAA), (1[->]3)-{beta}-D-Glucan (BG), and 16S rRNA gene sequencing to determine the circulating bacteriome. ResultsOf the 19 patients, 14 were in intensive care and 10 patients received mechanical ventilation. We found 8 patients with high EAA ([≥] 0.6) and about half of the patients had high serum BG levels which tended to be higher in later in the illness. Although only 1 patient had a positive blood culture, 18 of 19 patients were positive for 16S rRNA gene amplification. Proteobacteria was the most abundant phylum. The diversity of bacterial genera was decreased overtime. ConclusionsBacterial DNA and toxins were discovered in virtual all severely ill COVID-19 pneumonia patients. This raises a previously unrecognized concern for significant contribution of bacterial products in the pathogenesis of this disease

Interpretation of microscopic agglutination test for leptospirosis diagnosis and seroprevalence

Determination of antibody titer by microscopic agglutination test (MAT) has been used as a tool for leptospirosis diagnosis. Four fold or greater rise in antibody titers between acute and convalescent sera suggests recent Leptospira infection. In addition, results obtained by MAT have been used to predict infecting serovars. However, cross reactivity among various Leptospira serovars have been reported when patient sera were tested with a battery of Leptospira serovars. This study demonstrates cross- reactivity among several Leptospira serovars when MAT was performed on leptospirosis sera. The data support a role of MAT as a tool for diagnosis. However, for information on infecting serovars, Leptospira isolation and molecular identification should be performed.

Interpretation of microscopic agglutination test for leptospirosis diagnosis and seroprevalence

Determination of antibody titer by microscopic agglutination test (MAT) has been used as a tool for leptospirosis diagnosis. Four fold or greater rise in antibody titers between acute and convalescent sera suggests recent Leptospira infection. In addition, results obtained by MAT have been used to predict infecting serovars. However, cross reactivity among various Leptospira serovars have been reported when patient sera were tested with a battery of Leptospira serovars. This study demonstrates cross- reactivity among several Leptospira serovars when MAT was performed on leptospirosis sera. The data support a role of MAT as a tool for diagnosis. However, for information on infecting serovars, Leptospira isolation and molecular identification should be performed.

Cytokine levels in patients with chikungunya virus infection

OBJECTIVE@#To investigate cytokine profile in patients with chikungunya virus (CHIKV) infection.@*METHODS@#Twenty eight pairs of serum samples collected from CHIKV infected patients during the outbreak of chikungunya fever in South Thailand in 2008 were obtained. A multiple cytokine assay for detection of 17 cytokines was performed.@*RESULTS@#In the acute stage of CHIKV infection, the patients had significantly higher levels of interleukin-6, granulocyte colony-stimulating factor, granulocyte-macrophage colony-stimulating factor, monocyte chemotactic protein 1 and tumor necrosis factor alpha than the control (P<0.001, P=0.023, P=0.015, P<0.001 and P=0.024, respectively). When the disease developed to the recovery stage, the patients had significantly lower levels of interleukin-6, granulocyte-macrophage colony-stimulating factor, monocyte chemotactic protein 1 and macrophage inflammatory protein beta than in the acute stage (P<0.001).@*CONCLUSIONS@#This study provides additional information that these cytokines could play roles in pathogenesis of CHIKV infection and could be used as disease biomarkers or drug targets.