Virulence variation among epidemic and non-epidemic strains of Saint Louis encephalitis virus circulating in Argentina

Saint Louis encephalitis virus caused an outbreak of febrile illness and encephalitis cases in Córdoba, Argentina, in 2005. During this outbreak, the strain CbaAr-4005 was isolated from Culex quinquefasciatus mosquitoes. We hypothesised that this epidemic variant would be more virulent in a mouse model than two other non-epidemic strains (78V-6507 and CorAn-9275) isolated under different epidemiological conditions. To test this hypothesis, we performed a biological characterisation in a murine model, including mortality, morbidity and infection percentages and lethal infection indices using the three strains. Mice were separated into age groups (7, 10 and 21-day-old mice) and analysed after infection. The strain CbaAr-4005 was the most infective and lethal of the three variants, whereas the other two strains exhibited a decreasing mortality percentage with increasing animal age. The strain CbaAr-4005 produced the highest morbidity percentages and no significant differences among age groups were observed. The epidemic strain caused signs of illness in all inoculated animals and showed narrower ranges from the onset of symptoms than the other strains. CbaAr-4005 was the most virulent for Swiss albino mice. Our results highlight the importance of performing biological characterisations of arbovirus strains likely to be responsible for emerging or reemerging human diseases.

Morphology of the larvae, male genitalia and DNA sequences of Anopheles (Kerteszia) pholidotus (Diptera: Culicidae) from Colombia

Since 1984, Anopheles (Kerteszia) lepidotus has been considered a mosquito species that is involved in the transmission of malaria in Colombia, after having been incriminated as such with epidemiological evidence from a malaria outbreak in Cunday-Villarrica, Tolima. Subsequent morphological analyses of females captured in the same place and at the time of the outbreak showed that the species responsible for the transmission was not An. lepidotus, but rather Anopheles pholidotus. However, the associated morphological stages and DNA sequences of An. pholidotus from the foci of Cunday-Villarrica had not been analysed. Using samples that were caught recently from the outbreak region, the purpose of this study was to provide updated and additional information by analysing the morphology of female mosquitoes, the genitalia of male mosquitoes and fourth instar larvae of An. pholidotus, which was confirmed with DNA sequences of cytochrome oxidase I and rDNA internal transcribed spacer. A total of 1,596 adult females were collected in addition to 37 larval collections in bromeliads. Furthermore, 141 adult females, which were captured from the same area in the years 1981-1982, were analysed morphologically. Ninety-five DNA sequences were analysed for this study. Morphological and molecular analyses showed that the species present in this region corresponds to An. pholidotus. Given the absence of An. lepidotus, even in recent years, we consider that the species of mosquitoes that was previously incriminated as the malaria vector during the outbreak was indeed An. pholidotus, thus ending the controversy.

She’s a femme fatale: low-density larval development produces good disease vectors

Two hypotheses for how conditions for larval mosquitoes affect vectorial capacity make opposite predictions about the relationship of adult size and frequency of infection with vector-borne pathogens. Competition among larvae produces small adult females. The competition-susceptibility hypothesis postulates that small females are more susceptible to infection and predicts frequency of infection should decrease with size. The competition-longevity hypothesis postulates that small females have lower longevity and lower probability of becoming competent to transmit the pathogen and thus predicts frequency of infection should increase with size. We tested these hypotheses for Aedes aegypti in Rio de Janeiro, Brazil, during a dengue outbreak. In the laboratory, longevity increases with size, then decreases at the largest sizes. For field-collected females, generalised linear mixed model comparisons showed that a model with a linear increase of frequency of dengue with size produced the best Akaike’s information criterion with a correction for small sample sizes (AICc). Consensus prediction of three competing models indicated that frequency of infection increases monotonically with female size, consistent with the competition-longevity hypothesis. Site frequency of infection was not significantly related to site mean size of females. Thus, our data indicate that uncrowded, low competition conditions for larvae produce the females that are most likely to be important vectors of dengue. More generally, ecological conditions, particularly crowding and intraspecific competition among larvae, are likely to affect vector-borne pathogen transmission in nature, in this case via effects on longevity of resulting adults. Heterogeneity among individual vectors in likelihood of infection is a generally important outcome of ecological conditions impacting vectors as larvae.

Oseltamivir-resistant influenza A(H1N1)pdm2009 strains found in Brazil are endowed with permissive mutations, which compensate the loss of fitness imposed by antiviral resistance

The 2009 pandemic influenza A virus outbreak led to the systematic use of the neuraminidase (NA) inhibitor oseltamivir (OST). Consequently, OST-resistant strains, carrying the mutation H275Y, emerged in the years after the pandemics, with a prevalence of 1-2%. Currently, OST-resistant strains have been found in community settings, in untreated individuals. To spread in community settings, H275Y mutants must contain additional mutations, collectively called permissive mutations. We display the permissive mutations in NA of OST-resistant A(H1N1)pdm09 virus found in Brazilian community settings. The NAs from 2013 are phylogenetically distinct from those of 2012, indicating a tendency of positive selection of NAs with better fitness. Some previously predicted permissive mutations, such as V241I and N369K, found in different countries, were also detected in Brazil. Importantly, the change D344N, also predicted to compensate loss of fitness imposed by H275Y mutation, was found in Brazil, but not in other countries in 2013. Our results reinforce the notion that OST-resistant A(H1N1)pdm09 strains with compensatory mutations may arise in an independent fashion, with samples being identified in different states of Brazil and in different countries. Systematic circulation of these viral strains may jeopardise the use of the first line of anti-influenza drugs in the future.

Neutralising antibodies for Mayaro virus in Pantanal, Brazil

The Pantanal hosts diverse wildlife species and therefore is a hotspot for arbovirus studies in South America. A serosurvey for Mayaro virus (MAYV), eastern (EEEV), western (WEEV) and Venezuelan (VEEV) equine encephalitis viruses was conducted with 237 sheep, 87 free-ranging caimans and 748 equids, including 37 collected from a ranch where a neurologic disorder outbreak had been recently reported. Sera were tested for specific viral antibodies using plaque-reduction neutralisation test. From a total of 748 equids, of which 264 were immunised with vaccine composed of EEEV and WEEV and 484 had no history of immunisation, 10 (1.3%) were seropositive for MAYV and two (0.3%) for VEEV using criteria of a ≥ 4-fold antibody titre difference. Among the 484 equids without history of immunisation, 48 (9.9%) were seropositive for EEEV and four (0.8%) for WEEV using the same criteria. Among the sheep, five were sero- positive for equine encephalitis alphaviruses, with one (0.4%) for EEEV, one (0.4%) for WEEV and three (1.3%) for VEEV. Regarding free-ranging caimans, one (1.1%) and three (3.4%), respectively, had low titres for neutralising antibodies to VEEV and undetermined alphaviruses. The neurological disorder outbreak could not be linked to the alphaviruses tested. Our findings represent strong evidence that MAYV and all equine encephalitis alphaviruses circulated in the Pantanal.

Update on oral Chagas disease outbreaks in Venezuela: epidemiological, clinical and diagnostic approaches

Orally transmitted Chagas disease has become a matter of concern due to outbreaks reported in four Latin American countries. Although several mechanisms for orally transmitted Chagas disease transmission have been proposed, food and beverages contaminated with whole infected triatomines or their faeces, which contain metacyclic trypomastigotes of Trypanosoma cruzi, seems to be the primary vehicle. In 2007, the first recognised outbreak of orally transmitted Chagas disease occurred in Venezuela and largest recorded outbreak at that time. Since then, 10 outbreaks (four in Caracas) with 249 cases (73.5% children) and 4% mortality have occurred. The absence of contact with the vector and of traditional cutaneous and Romana’s signs, together with a florid spectrum of clinical manifestations during the acute phase, confuse the diagnosis of orally transmitted Chagas disease with other infectious diseases. The simultaneous detection of IgG and IgM by ELISA and the search for parasites in all individuals at risk have been valuable diagnostic tools for detecting acute cases. Follow-up studies regarding the microepidemics primarily affecting children has resulted in 70% infection persistence six years after anti-parasitic treatment. Panstrongylus geniculatushas been the incriminating vector in most cases. As a food-borne disease, this entity requires epidemiological, clinical, diagnostic and therapeutic approaches that differ from those approaches used for traditional direct or cutaneous vector transmission.

Molecular characterisation and disease severity of leptospirosis in Sri Lanka

Leptospirosis is a re-emerging zoonotic disease all over the world, important in tropical and subtropical areas. A majority of leptospirosis infected patients present as subclinical or mild disease while 5-10% may develop severe infection requiring hospitalisation and critical care. It is possible that several factors, such as the infecting serovar, level of leptospiraemia, host genetic factors and host immune response, may be important in predisposition towards severe disease. Different Leptospirastrains circulate in different geographical regions contributing to variable disease severity. Therefore, it is important to investigate the circulating strains at geographical locations during each outbreak for epidemiological studies and to support the clinical management of the patients. In this study immunochromatography, microscopic agglutination test and polymerase chain reaction were used to diagnose leptospirosis. Further restriction fragment length polymorphism and DNA sequencing methods were used to identify the circulating strains in two selected geographical regions of Sri Lanka.Leptospira interrogans, Leptospira borgpeterseniiandLeptospira kirschneristrains were identified to be circulating in western and southern provinces. L. interroganswas the predominant species circulating in western and southern provinces in 2013 and its presence was mainly associated with renal failure.

Detection of Oropouche virus segment S in patients and inCulex quinquefasciatus in the state of Mato Grosso, Brazil

This study aimed to investigate the circulation of Orthobunyavirus species in the state of Mato Grosso (MT) Brazil. During a dengue outbreak in 2011/2012, 529 serum samples were collected from patients with acute febrile illness with symptoms for up to five days and 387 pools of female Culex quinquefasciatuscaptured in 2013 were subjected to nested-reverse transcription-polymerase chain reaction for segment S of the Simbu serogroup followed by nucleotide sequencing and virus isolation in Vero cells. Patients (5/529; 0.9%) from Cuiabá (n = 3), Várzea Grande (n = 1) and Nova Mutum (n = 1) municipalities were positive for the S segment of Oropouche virus (OROV). Additionally, eight/387 Cx. quinquefasciatuspools were positive for the segment, with a minimum infection rate of 2.3. Phylogenetic analysis indicated that all the samples belong to the subgenotype Ia, presenting high homology with OROV strains obtained from humans and animals in the Brazilian Amazon. The present paper reports the first detection of an Orthobunyavirus, possibly OROV, in patients and in Cx. quinquefasciatus mosquitoes in MT. This finding reinforces the notion that arboviruses frequently reported in the Amazon Region circulate sporadically in MT during dengue outbreaks.

Yellow fever impact on brown howler monkeys (Alouatta guariba clamitans) in Argentina: a metamodelling approach based on population viability analysis and epidemiological dynamics

In South America, yellow fever (YF) is an established infectious disease that has been identified outside of its traditional endemic areas, affecting human and nonhuman primate (NHP) populations. In the epidemics that occurred in Argentina between 2007-2009, several outbreaks affecting humans and howler monkeys (Alouatta spp) were reported, highlighting the importance of this disease in the context of conservation medicine and public health policies. Considering the lack of information about YF dynamics in New World NHP, our main goal was to apply modelling tools to better understand YF transmission dynamics among endangered brown howler monkey (Alouatta guariba clamitans) populations in northeastern Argentina. Two complementary modelling tools were used to evaluate brown howler population dynamics in the presence of the disease: Vortex, a stochastic demographic simulation model, and Outbreak, a stochastic disease epidemiology simulation. The baseline model of YF disease epidemiology predicted a very high probability of population decline over the next 100 years. We believe the modelling approach discussed here is a reasonable description of the disease and its effects on the howler monkey population and can be useful to support evidence-based decision-making to guide actions at a regional level.

Selection of shrimp breeders free of white spot syndrome and infectious hypodermal and hematopoietic necrosis

The objective of this work was to select surviving breeders of Litopenaeus vannamei from white spot syndrome virus (WSSV) outbreak, adapted to local climatic conditions and negatively diagnosed for WSSV and infectious hypodermal and hematopoietic necrosis virus (IHHNV), and to evaluate if this strategy is a viable alternative for production in Santa Catarina, Brazil. A total of 800 males and 800 females were phenotypically selected in a farm pond. Nested-PCR analyses of 487 sexually mature females and 231 sexually mature males showed that 63% of the females and 55% of the males were infected with IHHNV. Animals free of IHHNV were tested for WSSV, and those considered double negative were used for breeding. The post-larvae produced were stocked in nine nursery tanks for analysis. From the 45 samples, with 50 post-larvae each, only two were positive for IHHNV and none for WSSV. Batches of larvae diagnosed free of virus by nested-PCR were sent to six farms. A comparative analysis was carried out in growth ponds, between local post-larvae and post-larvae from Northeast Brazil. Crabs (Chasmagnathus granulata), blue crabs (Callinectes sapidus), and sea hares (Aplysia brasiliana), which are possible vectors of these viruses, were also evaluated. The mean survival was 55% for local post-larvae against 23.4% for post-larvae from the Northeast. Sea hares showed prevalence of 50% and crabs of 67% of WSSV.

Mites fluctuation population on peach tree (Prunus persica (L.) Batsch) and in associated plants

Despite the importance of peach (Prunus persica (L.) Batsch) in Rio Grande do Sul, little is known about mites fluctuation population considered important to this crop. The objective of this study was to know the population diversity and fluctuation of mite species associated with Premier and Eldorado varieties in Roca Sales and Venâncio Aires counties, Rio Grande do Sul. The study was conducted from July 2008 to June 2009 when 15 plants were randomly chosen in each area. The plants were divided in quadrants and from each one a branch was chosen from which three leaves were removed: one collected in the apical region, another in the medium and the other in the basal region, totalizing 180 leaves/area. Five of the most abundant associated plants were collected monthly in enough amounts for the screening under the stereoscopic microscope during an hour. A total of 1,124 mites were found belonging to 14 families and 28 species. Tetranychus ludeni Zacher, 1913, Panonychus ulmi (Koch, 1836) and Mononychellus planki (McGregor, 1950) were the most abundant phytophagous mites, whereas Typhlodromalus aripo Deleon, 1967 and Phytoseiulus macropilis (Banks, 1904) the most common predatory mites. The period of one hour under stereoscopic microscope was enough to get a representative sample. In both places evaluated the ecologic indices were low, but little higherin Premier (H' 0.56; EqJ: 0.43) when compared to Eldorado (H' 0.53; EqJ 0.40). In Premier constant species were not observed and accessory only Brevipalpus phoenicis (Geijskes, 1939), T. ludeni and T. aripo. Higher abundance was observed in December and January and bigger amount in April. Already in Eldorado, T. ludeni and P. ulmi were constants. Greater abundance was observed in November and December, whereas grater richness in December and January. In both orchards were not found mites in buds. Tetranychus ludeni is the most abundant phytophagous mites with outbreak population in November, December and January and high predator diversity was observed on associated plants and on peach plants, indicating the existence of species mobility in peach orchard.

Genetic and phenotypic characterization of isolates of Pyricularia grisea from the rice cultivars epagri 108 and 109 in the State of Tocantins

An epidemic of rice (Oryza sativa) blast occurred on cultivars Epagri 108 and 109 in the municipalities of Lagoa da Confusão and Duerê in the State of Tocantins, during the rice-growing season 1998-99. DNA fingerprinting and virulence phenotype analysis were utilized to determine the diversity of Pyricularia grisea isolates collected from these cultivars in one epidemic year. Rep-PCR analysis of isolates was done by using two primer sequences from Pot2. Two distinct fingerprint groups or lineages were identified among 53 isolates collected from nine different commercial fields. The virulence pattern of isolates retrieved from these two cultivars was analyzed in artificial inoculation tests utilizing 32 genotypes in the greenhouse. A dendrogram constructed from virulence phenotype data showed a single group considering 77% similarity level. The predominant pathotype IB-45 was represented by 47 of the 53 isolates corresponding to 83%. Four other pathotypes (IB-1, IB-9, IB-13 and IB-41) were identified at random among the isolates from these cultivars. There was no relation between rep-PCR grouping and pathotypes. The results showed that the isolates of P. grisea recovered from cultivars Epagri108 and 109 in farmers' fields had narrow phenotypic and genetic diversity. The blast outbreak on these two cultivars one year after their introduction could be attributed to the new pathotype IB-45 or its increase, which was hitherto existing in low frequency.

Infecção por micobactérias de crescimento rápido após procedimentos videocirúrgicos - a hipótese do glutaraldeído

Between August 2006 and February 2007, in the state of Rio de Janeiro, Brazil, a massive outbreak of RGM infections after video laparoscopy was mainly associated to the recently described Mycobacterium massiliense species. All confirmed and probable cases reports described the use of high-level disinfection of medical devices by using 2% glutaraldehyde (2% GA) for 30 min before the surgical procedures. We investigated the susceptibility of the M. massiliense isolates recovered during the outbreak to high-level disinfection after 30 min, 1h, 6h and 10h of exposure to the commercial disinfectants. Reference strains for official mycobactericidal tests such as Mycobacterium abscessus, Mycobacterium bovis, Mycobacterium chelonae, Mycobacterium neoaurum and Mycobacterium smegmatis were included as controls. Although all the reference strains were eliminated in 30 min of exposure to 2% GA, we observed the recovery of all M. massiliense clinical isolates even after 10h of exposure. This study suggests that failures in high-level disinfection and the high tolerance of these M. massiliense clinical strains to the 2% GA were strongly associated to the magnitude of the outbreak.

Typing of avian pathogenic Escherichia coli strains by REP-PCR

In the present study the repetitive extragenic palindromic (REP) polymerase chain reaction (PCR) technique was used to establish the clonal variability of 49 avian Escherichia coli (APEC) strains isolated from different outbreak cases of septicemia (n=24), swollen head syndrome (n=14) and omphalitis (n=11). Thirty commensal strains isolated from poultry with no signs of these illnesses were used as control strains. The purified DNA of these strains produced electrophoretic profiles ranging from 0 to 15 bands with molecular sizes varying from 100 bp to 6.1 kb, allowing the grouping of the 79 strains into a dendrogram containing 49 REP-types. Although REP-PCR showed good discriminating power it was not able to group the strains either into specific pathogenic classes or to differentiate between pathogenic and non-pathogenic strains. On the contrary, we recently demonstrated that other techniques such as ERIC-PCR and isoenzyme profiles are appropriate to discriminate between commensal and APEC strains and also to group these strains into specific pathogenic classes. In conclusion, REP-PCR seems to be a technique neither efficient nor universal for APEC strains discrimination. However, the population clonal structure obtained with the use of REP-PCR must not be ignored particularly if one takes into account that the APEC pathogenic mechanisms are not completely understood yet.

Postweaning multisystemic wasting syndrome in farmed wild boars (Sus scrofa) in Rio Grande do Sul

This report concerns an outbreak of postweaning multisystemic wasting syndrome in wild boars from commercial herds in Rio Grande do Sul, Brazil. The main clinical signs were progressive wasting, respiratory distress, diarrhea, and ocular discharges. Enhanced lymph nodes and spleens, ascites, hydrothorax, and non collapsed lungs were the principal gross lesions. Varying degrees of lymphohistiocytic infiltrates in lymph nodes, spleen, and intestines were the most significant microscopic changes. Although a total of 26 animals had showed clinical and pathological findings consistent with the disease, only in 11 of them it was possible to confirm a PMWS diagnosis through the association of specific lesions in lymphoid tissues with the presence of the viral antigen detected by immunohistochemistry. These findings were similar to those reported in the swine disease.