A Rough Variant of Nontoxigenic Vibrio Cholerae O139 Imported From Vietnam

Purpose (stating the main purposes and research question)Vibrio cholerae is classified in O-antigen polysaccharide outer membrane properties where O1 and O139 are strains that cause pandemics and epidemics while non-O1/non-O139 usually cause mild disease. The dynamic evolution of Vibrio cholerae involves new virulence factors through horizontal gene transfer and formerly nontoxigenic serogroups are being reported as causing increasingly severe forms of human disease. Our purpose was to investigate serogroup and virulence factors in one imported isolate from Vietnam and compare these to virulence factors seen in different strains of Vibrio cholerae.MethodsWe have serotyped one isolate of imported Vibrio cholerae from Vietnam to Denmark and performed whole genome sequencing to identify known virulence genes. ResultsWe have identified virulence factors in our isolate that are recently discovered and one toxin, MakA, has not previously been reported from an Asian strain. The isolate was found to be a serogroup non-O1/non-O139 strain, however, sequence analysis gave a 96, 6% ID match of the wbfZ gene, stipulating our isolate to belong to either serogroup O22 or O139. ConclusionsLooking at the combination of virulence factors of the isolate we suggest the finding of a rough variant of a nontoxigenic Vibrio cholerae O139 rather than a O22 serogroup version. This is a rare finding in a clinical isolate.

TAL effectors with avirulence activity in African strains of Xanthomonas oryzae pv. oryzae

Background: Xanthomonas oryzae pv. oryzae causes bacterial leaf blight, a devastating disease of rice. Among the type-3 effectors secreted by Xanthomonas oryzae pv. oryzae to support pathogen virulence, the Transcription Activator-Like Effector (TALE) family plays a critical role. Some TALEs are major virulence factors that activate susceptibility (S) genes, overexpression of which contributes to disease development. Host incompatibility can result from TALE-induced expression of so-called executor (E) genes leading to a strong and rapid resistance response that blocks disease development. In that context, the TALE functions as an avirulence (Avr) factor. To date no such avirulence factors have been identified in African strains of Xanthomonas oryzae pv. oryzae. Results: With respect to the importance of TALEs in the Rice-Xoo pathosystem, we aimed at identifying those that may act as Avr factor within African Xoo. We screened 86 rice accessions, and identified 12 that were resistant to two African strains while being susceptible to a well-studied Asian strain. In a gain of function approach based on the introduction of each of the nine tal genes of the avirulent African strain MAI1 into the virulent Asian strain PXO99A, four were found to trigger resistance on specific rice accessions. Loss-of-function mutational analysis further demonstrated the avr activity of two of them, talD and talI, on the rice varieties IR64 and CT13432 respectively. Further analysis of TalI demonstrated the requirement of its activation domain for triggering resistance in CT13432. Resistance in 9 of the 12 rice accessions that were resistant against African Xoo specifically, including CT13432, could be suppressed or largely suppressed by trans-expression of the truncTALE tal2h, similarly to resistance conferred by the Xa1 gene which recognizes TALEs generally independently of their activation domain. Conclusion: We identified and characterized TalD and TalI as two African Xoo TALEs with avirulence activity on IR64 and CT13432 respectively. Resistance of CT13432 against African Xoo results from the combination of two mechanisms, one relying on the TalI-mediated induction of an unknown executor gene and the other on an Xa1-like gene or allele.

Leu-to-Phe substitution at prM146 decreases the growth ability of Zika virus and partially reduces its pathogenicity in mice

Zika virus (ZIKV) is a mosquito-borne flavivirus that causes febrile illness. The recent spread of ZIKV from Asia to the Americas via the Pacific region has revealed unprecedented features of ZIKV, including transplacental congenital infection causing microcephaly. Amino acid changes have been hypothesized to underlie the spread and novel features of American ZIKV strains; however, the relationship between genetic changes and the epidemic remains controversial. A comparison of the characteristics of a Southeast Asian strain (NIID123) and an American strain (PRVABC59) revealed that the latter had a higher replication ability in cultured cells and higher virulence in mice. In this study, we aimed to identify the genetic region of ZIKV responsible for these different characteristics using reverse genetics. A chimeric NIID123 strain in which the E protein was replaced with that of PRVABC59 showed a lower growth ability than the recombinant wild-type strain. Adaptation of the chimeric NIID123 to Vero cells induced a Phe-to-Leu amino acid substitution at position 146 of the prM protein; PRVABC59 also has Leu at this position. Leu at this position was found to be responsible for the viral replication ability and partially, for the pathogenicity in mouse testes.

ZIKV outbreak in Thiruvananthapuram, Kerala, India, 2021 –a primary report

The recent global outbreaks of the ZIKA Virus (ZIKV) reported in 85 countries and territories caused severe complications such as microcephaly among neonates and Guillain-Barre Syndrome among the older population. Recently, an outbreak of ZIKV was reported from Thiruvananthapuram, the capital city of Kerala, India with 66 cases being reported so far. We conducted an outbreak investigation and the primary findings are described here. A cluster of ZIKV cases from the Kadakampalli / Anamugham administrative wards of the Thiruvananthapuram Municipal Corporation area was reported where Kerala Institute of Medical Sciences (KIMS) is located. Later many ZIKV cases were reported from other wards of the city. The density of known Aedes vectors was high in this region of the metropolitan city. Aedes albopictus, Aedes aegypti and Aedes vittatus collected from the focal area of the outbreak were found to be naturally infected with ZIKV. Male specimens of Ae. albopictus were naturally infected, indicating trans-ovarian transmission of the virus. This is first report of incrimination of Ae. albopictus and Ae. vittatus in ZIKV transmission from India. The virus was characterized and the partial sequences clustered with the Asian strain of ZIKV reported from India. The NS5 sequences of human and Ae. albopictus pools from Thiruvananthapuram were 100% similar indicating an ongoing active ZIKV transmission. The state health authorities were sensitized and appropriate containment and vector control measures have been initiated to contain the outbreak. This report underscores the importance of continued human and vector surveillance as well as genomic sequencing to understand the virus evolution and implications on public health.

Molecular Identification of Sweet potato virus C on Sweetpotato in Bali, Indonesia

A survey was conducted in several sweet potato cultivations in Bali Province. Survey found that many plants exhibited potyvirus symptom, such as chlorosis blotches. This study was to determine disease incidence, detection and identification of the virus causing these symptoms on sweet potato plants in Bali. Samples were collected by purposive sampling of 10 plants from each location in Bali (Denpasar, Gianyar, Badung, Buleleng, Tabanan, Klungkung, Karangasem, Jembrana, Bangli). Disease insidence was observed based on viral symptoms in the field. Identification of nucleic acids was done using Potyvirus universal primer and DNA sequencing. Disease incidence in Bangli, Buleleng, and Denpasar Regencies was > 50%. RT-PCR and CiFor/CiRev Potyvirus universal primers successfully amplified ± 700 bp of CI genes from all samples from Bangli, while samples from 8 other districts were not amplified using the same primers. The SPVC isolate of sweet potato showed nucleotide and amino acid homology similarities with the sweet potato isolate from East Timor (MF572066), 96.8% and 97.4%, respectively and these were referred to the "Asian" strain. This indicates that SPVC has spread in East Java and Bali.

Resistance to Cucumber Green Mottle Mosaic Virus in Cucumis melo

Cucumber green mottle mosaic virus (CGMMV) is a severe threat to melon production worldwide. At present, there are no cultivars available on the market which show an effective resistance or tolerance to CGMMV infection; only wild Cucumis species were reported as resistant. Germplasm accessions of Cucumis melo, as well as C. anguria, C. ficifolius, C. myriocarpus and C. metuliferus, were mechanically infected with isolates belonging to the European and Asian strain of CGMMV and screened for resistance by scoring symptom severity and comparing the accumulation of virus by qRT-PCR. The wild species C. anguria and C. ficifolius showed no symptoms and did not accumulate CGGMV following inoculation, while C. metuliferus was highly susceptible to the isolates of both strains of CGMMV. The virus accumulated also in C. myriocarpus and the European isolate produced symptoms, but the Asian isolate did not. Thirty C. melo accessions were susceptible to CGMMV. An isolate-dependent expression of symptoms was observed in 16 melon accessions: they showed mild and severe symptoms at 14 and 21 days after inoculation with the European and Asian isolate, respectively. Freeman’s Cucumber showed few or no symptoms following inoculation with the isolate of either CGMMV strain. This particular accession also showed reduced virus accumulation, whereas most other tested germplasm accessions showed significantly higher viral loads and, therefore, may well be a candidate for breeding programs aiming to reduce the losses produced by CGMMV with resistant commercial melon cultivars.

Lymantria dispar (gypsy moth).

The gypsy moth is likely to ultimately occupy virtually all portions of the temperate world where oaks and other suitable host plants occur. Consequently, the northern hemisphere is more at risk for establishment than the southern hemisphere though some suitable hosts do occur in these areas. The gypsy moth is apparently not able to persist in very cold (e.g. Finland) or warm (subtropical to tropical) regions. The gypsy moth is a 'proven' invader. The broad range of host plants that it utilizes (Liebhold et al. 1995), along with its high reproductive rate combine to make this insect a very successful invader of many types of forest and urban landscapes. Another characteristic that contributes to the gypsy moth's invasiveness is its propensity to be transported on human-made objects (e.g., egg masses can be laid on vehicles, logs, etc.). Perhaps the greatest limitation this species has as an invader is that females (of the European strain) are incapable of flight and this limits its rate of unassisted range expansion. However, as females of the Asian strain are capable of flight and all strains can lay their eggs on human-made objects, established populations are nevertheless able to spread.

Molecular characterization and the mutation pattern of SARS-CoV-2 during first and second wave outbreaks in Hiroshima, Japan

Background In this study, we performed molecular characterization of SARS-CoV-2 strains in Hiroshima and its mutation pattern between the first and second waves of the outbreak. Method A total of 55 nasal swab samples from the first wave in Hiroshima and 13 from the second wave were examined quantitatively by RT-qPCR and qualitatively by nested PCR using specific primers. Four samples from each wave underwent next-generation sequencing and phylogenetic tree analysis including controls and all sequences retrieved in Japan from GISAID and GenBank. Subsequently, mutations were examined. Results Viral load ranged 7.85 × 101−1.42 × 108 copies/ml. Of 68 samples, one was Asian type-O, 65 were European type-GR, and 2 were undetectable. Phylogenetic tree analysis indicated that Japan was infected with various Asian strains (L, S, V, O) from January through April. By second week of March, European strains (G, GH, GR) had appeared, and GR strains became predominant after mid-March. The first case in Hiroshima was classified as Asian strain O, and the rest were GR strains. Then, second wave of GR strains appeared independently with 11–15 base mutations. Comparing the first- and second-wave GR strains, mutation rate was 1.17–1.36 × 10−3 base substitutions per site per year; in addition, amino acid changes occurred at S1361P and P3371S in ORF1a, A314V in ORF1b, and P151L in N. All seven GR strains were D614G variants with R202K and G203R mutations in N. A single-nucleotide insertion in ORF8 that causes a defect in ORF8 protein was found in one isolate (S66) from the second wave. Conclusion Our findings reveal the evolutionary hierarchy of SARS-CoV-2 in Japan. The predominant D614G variants and a new form of ORF8 deletion in Hiroshima provide the clue for role of viral factor in local outbreaks of SARS-CoV-2.

Infection by Zika Vírus in human cells alters the expression profile of miRNA-15 and activation of apoptotic caspases

Objective: Evaluate the miRNA-15 expression profile involved in cellular apoptotic regulation factors. Methodology: We used the H818308 Asian strain of ZIKV without neurological damage. The inoculations occurred in human embryonic kidney cells (HEK-293). After inoculation, samples were extracted for RT-qPCR quantification of viral RNA and miR-15. The level of activation of caspases 1, 3/7 and 8 of cells was performed using chemofluorescence. Results: The ZIKV infection alters the expression of genes and their regulators, affecting several cellular physiological processes such as apoptosis. Conclusion: Therefore, it is important to emphasize that renal progenitor cells (HEK-293) are susceptible to VZIK infection. The genetic deregulation resulting from infection directly affects important cellular processes such as apoptosis from the disordered miRNA-15 expression during the infection period.