Phylogeography and evolutionary history of hepatitis E virus genotype 3 in Argentina.

BACKGROUND: Few studies about the evolutionary history of the hepatitis E virus (HEV) have been conducted. The aim of our work was to investigate and make inferences about the origin and routes of dispersion of HEV-3 in Argentina. METHODS: Phylogenetic, coalescent and phylogeographic analyses were performed using a 322-bp ORF2 genomic fragment of all HEV-3 sequences with known date and place of isolation published at GenBank until May 2018 (n=926), including 16 Argentinian sequences (isolated from pigs, water and humans). RESULTS: Phylogenetic analysis revealed two clades within HEV-3: abchij and efg. All Argentinian samples were grouped intermingled within clade 3abchij. The coalescent analysis showed that the most recent common ancestor for the clade 3abchij would have existed around the year 1967 (95% highest posterior density (HPD): 1963-1970). The estimated substitution rate was 1.01×10-2 (95%HPD: 9.3×10-3-1.09×10-2) substitutions/site/y, comparable with the rate previously described. The phylogeographic approach revealed a correspondence between phylogeny and place of origin for Argentinian samples, suggesting many HEV introductions in the country, probably from Europe and Japan. CONCLUSIONS: This is the first evolutionary inference of HEV-3 that includes Argentinian strains, showing the circulation of many HEV-3 subtypes, obtained from different sources and places, with recent diversification processes. ACCESSION NUMBERS: [KX812460], [KX812461], [KX812462], [KX812465], [KX812466], [KX812467], [KX812468], [KX812469].

Genotypes and phylogenetic analysis of adenovirus in children with respiratory infection in Buenos Aires, Argentina (2000-2018).

Human adenoviruses (HAdV) are one of the most frequent causes of respiratory infections around the world, causing mild to severe disease. In Argentina, many studies focused on the association of HAdV respiratory infection with severe disease and fatal outcomes leading to the discovery in 1984 of a genomic variant 7h associated with high fatality. Although several molecular studies reported the presence of at least 4 HAdV species (B, C, D and E) in Argentina, few sequences were available in the databases. In this study, sequences from the hexon gene region were obtained from 141 patients as a first approach to assess the genetic diversity of HAdVs circulating in Buenos Aires, Argentina. Phylogenetic analysis of these sequences and others recovered from public databases confirmed the circulation of the four above-mentioned species represented by 11 genotypes, with predominance in species B and C and shifts in their proportion in the studied period (2000 to 2018). The variants detected in Argentina, for most of the genotypes, were similar to those already described in other countries. However, uncommon lineages belonging to genotypes C2, C5 and E4 were detected, which might indicate the circulation of local variants and will deserve further studies of whole-genome sequences.

Longitudinal characterization of HIV-1 pol-gene in treatment-naïve men-who-have-sex-with-men from acute to chronic infection stages.

HIV-1 is characterized by its ability to mutate and recombine even at polymerase (pol) gene. However, pol-gene diversity is limited due to functional constraints. The aim of this study was to characterize longitudinally, by next-generation sequencing (NGS), HIV-1 variants based on pol-gene sequences, at intra- and inter-host level, from acute/early to chronic stages of infection, in the absence of antiretroviral therapy. Ten men who have sex with men (MSM) were recruited during primary infection and yearly followed for five years. Even after a maximum of a five-year follow-up period, the phylogenetic analysis of HIV-1 pol-gene sequences showed a host-defined structured pattern, with a predominance of purifying selection forces during the follow-up. MSM had been acutely infected by different HIV-1 variants mainly ascribed to pure subtype B, or BF recombinant variants and showed different genetic mosaicism patterns that last until the chronic stage, representing a major challenge for prevention strategies.

In vivo drug resistance mutation dynamics from the early to chronic stage of infection in antiretroviral-therapy-naïve HIV-infected men who have sex with men.

Human immunodeficiency virus type 1 (HIV) primary drug resistance mutations (DRMs) influence the long-term therapeutic effects of antiretroviral treatment (ART). Drug-resistance genotyping based on polymerase gene sequences obtained by next-generation sequencing (NGS) was performed using samples from 10 ART-naïve HIV-infected men who have sex with men (MSM; P1-P10) from the acute/early to chronic stage of infection. Three of the 10 subjects exhibited the presence of major (abundance, ≥ 20%) viral populations carrying DRM at early/acute stage that later, at the chronic stage, dropped drastically (V106M) or remained highly abundant (E138A). Four individuals exhibited additional DRMs (M46I/L; I47A; I54M, L100V) as HIV minority populations (abundance, 2-20%) that emerged during the chronic stage but ephemerally.

Molecular epidemiology and phylogenetic analysis of human papillomavirus infection in women with cervical lesions and cancer from the coastal region of Ecuador / Epidemiología molecular y análisis filogenético de la infección por el virus del papiloma humano en mujeres con lesiones cervicales y cáncer en la región litoral del Ecuador

The aim of the present study was to gather information regarding the molecular epidemiology of Human papillomavirus (HPV) and related risk factors in a group of women with low- and high-grade cervical lesions and cancer from the coastal region of Ecuador. In addition, we studied the evolution of HPV variants from the most prevalent types and provided a temporal framework for their emergence, which may help to trace the source of dissemination within the region. We analyzed 166 samples, including 57 CIN1, 95 CIN2/3 and 14 cancer cases. HPV detection and typing was done by PCR-sequencing (MY09/MY11). HPV variants and estimation of the time to most recent common ancestor (tMRCA) was assessed through phylogeny and coalescence analysis. HPV DNA was found in 54.4% of CIN1, 74.7% of CIN2/3 and 78.6% of cancer samples. HPV16 (38.9%) and HPV58 (19.5%) were the most prevalent types. Risk factors for the development of cervical lesions/cancer were the following: three or more pregnancies (OR = 4.3), HPV infection (OR = 3.7 for high-risk types; OR = 3.5 for HPV16), among others. With regard to HPV evolution, HPV16 isolates belonged to lineages A (69%) and D (31%) whereas HPV58 isolates belonged only to lineage A. The period of emergence of HPV16 was in association with human populations (tMRCA = 91 052 years for HPV16A and 27000 years for HPV16D), whereas HPV58A preceded Homo sapiens evolution (322 257 years). This study provides novel data on HPV epidemiology and evolution in Ecuador, which will be fundamental in the vaccine era.

El objetivo del presente estudio fue aportar información sobre la epidemiología molecular del virus del papiloma humano (human papillomavirus [HPV]) y los factores de riesgo asociados al desarrollo de lesiones cervicales y cáncer en mujeres de la costa del Ecuador. Además, se estudiaron la evolución de las variantes de los HPV más prevalentes y el marco temporal de su emergencia, para ayudar a rastrear la fuente de dispersión en la región. Se analizaron 166 muestras, incluyendo 57 y 95 casos de neoplasia intraepitelial cervical tipo 1 (CIN1) y tipo 2/3 (CIN2/3), respectivamente, y 14 de casos de cáncer. La detección/tipificación de HPV se realizó por PCR-secuenciación (MY09/MY11). La caracterización de variantes y la datación del ancestro común más reciente (tMRCA) se realizaron mediante filogenia y coalescencia. Se encontró ADN de HPV en el 54,4% de las muestras de CIN1, el 74,7% de las muestras de CIN2/3 y el 78,6% de las muestras de cáncer. Los tipos HPV16 (38,9%) y HPV58 (19,5%) fueron los más frecuentes. Los factores de riesgo para el desarrollo de lesiones cervicales/cáncer fueron 3 o más embarazos (OR = 4,3) e infección por HPV (O = 3,7 para HPV de alto riesgo, OR = 3,5 para HPV16), entre otros. En cuanto a la evolución viral, los aislados del HPV16 pertenecían a los linajes A (69%) y D (31%), mientras que los aislados del HPV58 pertenecían únicamente al linaje A. El período de emergencia del HPV16 estuvo asociado a poblaciones humanas (tMRCA = 91.052 años para HPV16Ay 27.000 para HPV16D), mientras que el del HPV58A precedió a la evolución de Homo sapiens (322.257 años). Este estudio proporciona datos novedosos sobre la epidemiología y la evolución del HPV en Ecuador, los cuales serán fundamentales en la era de la vacuna.

Molecular epidemiology and phylogenetic analysis of human papillomavirus infection in women with cervical lesions and cancer from the coastal region of Ecuador.

The aim of the present study was to gather information regarding the molecular epidemiology of Human papillomavirus (HPV) and related risk factors in a group of women with low- and high-grade cervical lesions and cancer from the coastal region of Ecuador. In addition, we studied the evolution of HPV variants from the most prevalent types and provided a temporal framework for their emergence, which may help to trace the source of dissemination within the region. We analyzed 166 samples, including 57 CIN1, 95 CIN2/3 and 14 cancer cases. HPV detection and typing was done by PCR-sequencing (MY09/MY11). HPV variants and estimation of the time to most recent common ancestor (tMRCA) was assessed through phylogeny and coalescence analysis. HPV DNA was found in 54.4% of CIN1, 74.7% of CIN2/3 and 78.6% of cancer samples. HPV16 (38.9%) and HPV58 (19.5%) were the most prevalent types. Risk factors for the development of cervical lesions/cancer were the following: three or more pregnancies (OR=4.3), HPV infection (OR=3.7 for high-risk types; OR=3.5 for HPV16), among others. With regard to HPV evolution, HPV16 isolates belonged to lineages A (69%) and D (31%) whereas HPV58 isolates belonged only to lineage A. The period of emergence of HPV16 was in association with human populations (tMRCA=91052 years for HPV16A and 27000 years for HPV16D), whereas HPV58A preceded Homo sapiens evolution (322257 years). This study provides novel data on HPV epidemiology and evolution in Ecuador, which will be fundamental in the vaccine era.

Polymorphisms associated with resistance to protease inhibitors in naïve patients infected with hepatitis C virus genotype 1 in Argentina: Low prevalence of Q80K.

Incorporation of direct acting antivirals (DAA) in the treatment of Hepatitis C Virus (HCV) significantly increases sustained virologic response rates. However, despite the greater potency offered by these antivirals, drug resistance plays a key role in patients with failure to DAA. Nevertheless, there is no information about the prevalence of resistance-associated substitutions (RASs) in Argentina. The aim of this study was to analyze HCV variants resistant to protease inhibitors (PI) in naïve patients infected with HCV genotype 1 from Argentina. In this retrospective cross-sectional study, 103 patients infected with HCV-1 were included. Eighteen positions related with RASs were analyzed by Sanger at baseline and phylogenetic analysis was performed to determine the diversification of this samples. The analyzed RASs were present in 38 out of 103 patients (36.9%) infected with HCV-1. Patients infected with subtype HCV-1b had higher prevalence of baseline RASs than patients infected with HCV-1a [51.6% vs. 12.8%, respectively (p<0.001)]. The Q80K polymorphism was not found in HCV-1a samples, even when 51% of them belonged to cluster 1, which is associated with a high frequency of Q80K. Phylogenetic analysis showed that Argentinean samples were intermingled with sequences from other geographic regions. RASs to PI were highly prevalent and subtype dependent in treatment-naïve Argentinean patients. Surprisingly, Q80K polymorphism was not detected in our study population. The phylogenetic analysis showed no relationship between our samples and other samples from Brazil which also present a low prevalence of Q80K. This study supports the need for surveillance of resistance in patients who will be treated with DAA in each particular country since the observed RASs have very different prevalence worldwide.

NS3 genomic sequencing and phylogenetic analysis as alternative to a commercially available assay to reliably determine hepatitis C virus subtypes 1a and 1b.

OBJECTIVE: To evaluate the use of hepatitis C virus (HCV) NS3 sequencing as alternative to the comercially available Versant HCV 2.0 reverse hybridization line-probe assay (LiPA 2.0) to determine HCV genotype 1 (HCV-1) subtypes. PATIENTS AND METHODS: A cohort of 104 patients infected by HCV-1 according to LiPA 2.0 was analyzed in a cross-sectional study conducted in patients seen from January 2012 to June 2016 at an outpatient clinic in Buenos Aires, Argentina. RESULTS: The samples were included within well supported subtype clades: 64 with HCV-1b and 39 with HCV-1a infection. Twenty of the HCV-1a infected patientes were included in a supported sub-clade "1" and 19 individuals were among the basal sub-clade "2". LiPA 2.0 failed to subtype HCV-1 in 20 (19.2%) individuals. Subtype classification determined by NS3 direct sequencing showed that 2/18 (11.1%) of the HCV-1a-infected patients as determined by LiPA 2.0 were in fact infected by HCV-1b. Of the HCV-1b-infected according to LiPA 2.0, 10/66 (15.2%) patients showed HCV-1a infection according to NS3 sequencing. Overall misclassification was 14.3% (κ-index for the concordance with NS3 sequencing = 0.635). One (1%) patient was erroneously genotyped as HCV-1 and was revealed as HCV genotype 4 infection. CONCLUSIONS: Genomic sequencing of the HCV NS3 region represents an adequate alternative since it provides reliable genetic information. It even distinguishes between HCV-1a clades related to resistance-associated substitutions to HCV protease inhibitors, it provides reliable genetic information for genotyping/subgenotyping and simultaneously allows to determine the presence of resistance-associated substitutions to currently recommended DAAs.

Inter and intrapatient evolution of hepatitis C virus.

Hepatitis C virus (HCV) has a short replication time, high mutation rates and large population sizes, all of which make it an excellent experimental model for evolution studies, because evolution can be visualized in real-time. In this review, we discuss the implications to study HCV evolution at the interpatient and intrapatient levels of infection. The HCV interpatient dynamics is relatively slow, because the generation time is generally long. Then, at population level, the HCV diversity originated by the high mutation and replication rates is modulated by the bottleneck at transmission. Thus, when the virus is transmitted to other hosts, viral diversity is reduced as a result of the founder effect. On the other hand, during intrapatient infection, HCV evolves rapidly, resulting in quasispecies. Accumulated evidence suggests that this quasispecies composition of the HCV population within the same individual may allow the virus to evade the immune response or escape treatment, leading to chronic infection. Thus, a better understanding of the complexities underlying the molecular evolution of HCV in natural populations is needed before accurate predictions of viral evolution can be made. In summary, HCV evolves both within and among patients. Consequently, HCV evolution should be studied at both levels in order to better understand the natural history of the virus and its potential implications in epidemiology, outcome of infection and progression of liver disease.

Rich bacterial assemblages from Maritime Antarctica (Potter Cove, South Shetlands) reveal several kinds of endemic and undescribed phylotypes.

Bacterial richness in maritime Antarctica has been poorly described to date. Phylogenetic affiliation of seawater free-living microbial assemblages was studied from three locations near the Argentinean Jubany Station during two Antarctic summers. Sixty 16S RNA cloned sequences were phylogenetically affiliated to Alphaproteobacteria (30/60 clones), Gammaproteobacteria(19/60 clones), Betaproteobacteria and Cytophaga-Flavobacteriia-Bacteroides (CFB), which were (2/60) and (3/60) respectively. Furthermore, six out of 60 clones could not be classified. Both, Alphaproteobacteria and Gammaproteobacteria, showed several endemic and previously undescribed sequences. Moreover, the absence of Cyanobacteria sequences in our samples is remarkable. In conclusion, we are reporting a rich sequence assemblage composed of widely divergent isolates among themselves and distant from the most closely related sequences currently deposited in data banks.

Rich bacterial assemblages from Maritime Antarctica (Potter Cove, South Shetlands) reveal several kinds of endemic and undescribed phylotypes / Una rica colección de bacterias marinas antárticas (Caleta Potter, islas Shetlands del Sur) revela diversos filotipos endémicos y previamente no descritos

.

Bacterial richness in maritime Antarctica has been poorly described to date. Phylogenetic affiliation of seawater free-living microbial assemblages was studied from three locations near the Argentinean Jubany Station during two Antarctic summers. Sixty 16S RNA cloned sequences were phylogenetically affiliated to Alphaproteobacteria (30/60 clones), Gammaproteobacteria(19/60 clones), Betaproteobacteria and Cytophaga-Flavobacteriia- Bacteroides (CFB), which were (2/60) and (3/60) respectively. Furthermore, six out of 60 clones could not be classified. Both, Alphaproteobacteria and Gammaproteobacteria, showed several endemic and previously undescribed sequences. Moreover, the absence of Cyanobacteria sequences in our samples is remarkable. In conclusion, we are reporting a rich sequence assemblage composed of widely divergent isolates among themselves and distant from the most closely related sequences currently deposited in data banks.

Rich bacterial assemblages from Maritime Antarctica (Potter Cove, South Shetlands) reveal several kinds of endemic and undescribed phylotypes / Una rica colección de bacterias marinas antárticas (Caleta Potter, islas Shetlands del Sur) revela diversos filotipos endémicos y previamente no descritos

.(AU)

Bacterial richness in maritime Antarctica has been poorly described to date. Phylogenetic affiliation of seawater free-living microbial assemblages was studied from three locations near the Argentinean Jubany Station during two Antarctic summers. Sixty 16S RNA cloned sequences were phylogenetically affiliated to Alphaproteobacteria (30/60 clones), Gammaproteobacteria(19/60 clones), Betaproteobacteria and Cytophaga-Flavobacteriia- Bacteroides (CFB), which were (2/60) and (3/60) respectively. Furthermore, six out of 60 clones could not be classified. Both, Alphaproteobacteria and Gammaproteobacteria, showed several endemic and previously undescribed sequences. Moreover, the absence of Cyanobacteria sequences in our samples is remarkable. In conclusion, we are reporting a rich sequence assemblage composed of widely divergent isolates among themselves and distant from the most closely related sequences currently deposited in data banks.(AU)

Rich bacterial assemblages from Maritime Antarctica (Potter Cove, South Shetlands) reveal several kinds of endemic and undescribed phylotypes. / Rich bacterial assemblages from Maritime Antarctica (Potter Cove, South Shetlands) reveal several kinds of endemic and undescribed phylotypes.

Bacterial richness in maritime Antarctica has been poorly described to date. Phylogenetic affiliation of seawater free-living microbial assemblages was studied from three locations near the Argentinean Jubany Station during two Antarctic summers. Sixty 16S RNA cloned sequences were phylogenetically affiliated to Alphaproteobacteria (30/60 clones), Gammaproteobacteria(19/60 clones), Betaproteobacteria and Cytophaga-Flavobacteriia-Bacteroides (CFB), which were (2/60) and (3/60) respectively. Furthermore, six out of 60 clones could not be classified. Both, Alphaproteobacteria and Gammaproteobacteria, showed several endemic and previously undescribed sequences. Moreover, the absence of Cyanobacteria sequences in our samples is remarkable. In conclusion, we are reporting a rich sequence assemblage composed of widely divergent isolates among themselves and distant from the most closely related sequences currently deposited in data banks.

Hepatitis C virus diversification in Argentina: comparative analysis between the large city of Buenos Aires and the small rural town of O'Brien.

BACKGROUND: The estimated prevalence of HCV infection in Argentina is around 2%. However, higher rates of infection have been described in population studies of small urban and rural communities. The aim of this work was to compare the origin and diversification of HCV-1b in samples from two different epidemiological scenarios: Buenos Aires, a large cosmopolitan city, and O'Brien, a small rural town with a high prevalence of HCV infection. PATIENTS AND METHODS: The E1/E2 and NS5B regions of the viral genome from 83 patients infected with HCV-1b were sequenced. Phylogenetic analysis and Bayesian Coalescent methods were used to study the origin and diversification of HCV-1b in both patient populations. RESULTS: Samples from Buenos Aires showed a polyphyletic behavior with a tMRCA around 1887-1900 and a time of spread of infection approximately 60 years ago. In contrast, samples from ÓBrien showed a monophyletic behavior with a tMRCA around 1950-1960 and a time of spread of infection more recent than in Buenos Aires, around 20-30 years ago. CONCLUSION: Phylogenetic and coalescence analysis revealed a different behavior in the epidemiological histories of Buenos Aires and ÓBrien. HCV infection in Buenos Aires shows a polyphyletic behavior and an exponential growth in two phases, whereas that in O'Brien shows a monophyletic cluster and an exponential growth in one single step with a more recent tMRCA. The polyphyletic origin and the probability of encountering susceptible individuals in a large cosmopolitan city like Buenos Aires are in agreement with a longer period of expansion. In contrast, in less populated areas such as O'Brien, the chances of HCV transmission are strongly restricted. Furthermore, the monophyletic character and the most recent time of emergence suggest that different HCV-1b ancestors (variants) that were in expansion in Buenos Aires had the opportunity to colonize and expand in O'Brien.

Molecular survey of hepatitis C virus in the touristic city of Mar del Plata, Argentina.

The global epidemiology of hepatitis C virus (HCV) may be roughly described by two groups of genotypes: the worldwide distributed ones (subtypes 1a, 1b, 2a and 3a, among others) and the endemic ones (subtypes 4a, 5a, 6a, among others). Epidemiological and population dynamic studies of the worldwide distributed genotypes have shown that subtypes 1a and 3a are common among intravenous drug users (IDUs) and that they are also in expansion in some countries. The molecular survey of HCV provides some clues about the epidemiological status of the infections in a local scale and the phylogenetic and demographic reconstruction analyses complement this study by inferring whether the infections of certain subtypes are in a steady state or expanding. Here, a molecular survey of the HCV variants that circulate in the touristic city of Mar del Plata (Buenos Aires, Argentina) was performed in samples obtained from 42 patients. The subtypes detected were 1a (32 patients), 3a (8 patients) and 1b (2 patients). The demographic history of subtype 1a inferred using the sequence data showed an exponential growth in the 1990's. The period of viral expansion was delayed compared with that observed for the same genotype in other countries where the transmission was associated with IDUs. Also, the phylogeographic analysis of HCV-1a showed a statistically significant association between the location of the samples and the phylogeny, which may be the result of the local transmission of HCV in the city. The molecular analysis helped in the description of the complex epidemiological context of a touristic city, and pointed out that some sanitary measures should be taken in order to reduce the transmission of HCV (and maybe of HIV) among IDUs.

Phylodynamics of hepatitis C virus subtype 2c in the province of Córdoba, Argentina.

The Hepatitis C Virus Genotype 2 subtype 2c (HCV-2c) is detected as a low prevalence subtype in many countries, except in Southern Europe and Western Africa. The current epidemiology of HCV in Argentina, a low-prevalence country, shows the expected low prevalence for this subtype. However, this subtype is the most prevalent in the central province of Córdoba. Cruz del Eje (CdE), a small rural city of this province, shows a prevalence for HCV infections of 5%, being 90% of the samples classified as HCV-2c. In other locations of Córdoba Province (OLC) with lower prevalence for HCV, HCV-2c was recorded in about 50% of the samples. The phylogenetic analysis of samples from Córdoba Province consistently conformed a monophyletic group with HCV-2c sequences from all the countries where HCV-2c has been sequenced. The phylogeographic analysis showed an overall association between geographical traits and phylogeny, being these associations significant (α = 0.05) for Italy, France, Argentina (places other than Córdoba), Martinique, CdE and OLC. The coalescence analysis for samples from CdE, OLC and France yielded a Time for the Most Common Recent Ancestor of about 140 years, whereas its demographic reconstruction showed a "lag" phase in the viral population until 1880 and then an exponential growth until 1940. These results were also obtained when each geographical area was analyzed separately, suggesting that HCV-2c came into Córdoba province during the migration process, mainly from Europe, which is compatible with the history of Argentina of the early 20th century. This also suggests that the spread of HCV-2c occurred in Europe and South America almost simultaneously, possibly as a result of the advances in medicine technology of the first half of the 20th century.