Hematúria macroscópica em equinos associada à infecção por Leptospira interrogans / Macroscopic haematuria in horses associated with Leptospira interrogans infection

Hematúria é uma grave manifestação clínica de doença do sistema urinário, ocorrendo sob as formas micro ou macroscópica. Neste artigo relatam-se dois casos de hematúria macroscópica associada à infecção por Leptospira interrogans sorogrupo Canicola. O exame clínico inicial revelou hematúria macroscópica, taquicardia, taquipneia, febre, elevação do tempo de perfusão capilar, hipomotilidade intestinal, além de icterícia da mucosa oral. Leucocitose, proteinúria, glicosúria, piúria e azotemia foram achados comuns aos dois casos. Teste de Soroaglutinação Microscópica foi realizado para titulação de anticorpos contra Leptospira interrogans. Tratamento incluiu medidas terapêuticas de suporte (fluidoterapia), controle da hematúria e antibioticoterapia. Sete dias após manifestação dos sinais clínicos iniciais, ambos animais receberam alta hospitalar após remissão dos sinais clínicos.

Haematuria is a serious clinical manifestation of urinary system disease, occurring in micro or macroscopic forms. In this article two cases of macroscopic haematuria associated with Leptospira interrogans serogroup Canicolainfection are related. The initial clinical examination revealed macroscopic haematuria, tachycardia, tachypnea, fever, increased capillary perfusion time, intestinal hypomotility, in addition to jaundice of the oral mucosa. Leukocytosis, proteinuria, glycosuria, pyuria and azotemia were common findings in both cases. Microscopic serum agglutination test was performed for titration of antibodies against Leptospira interrogans. Treatment included supportive therapeutic measures (fluid therapy), hematuria control and antibiotic therapy. Seven days after the manifestation of the initial clinical signs, both animals were discharged from the hospital without complications.

Genotipificación y evaluación de la dinámica de infección de un aislamiento colombiano de Leptospira santarosai en el modelo experimental en hámster / Genotyping and evaluation of infection dynamics in a Colombian isolate of Leptospira santarosai in hamster as an experimental model

Introducción. Es necesario desarrollar modelos de estudio de la leptospirosis. Objetivo. Genotipificar un aislamiento de Leptospira proveniente de una persona con síndrome de Weil y evaluar, con el modelo experimental en Mesocricetus auratus , su dinámica de infección. Materiales y métodos. Se hizo la genotipificación por análisis de las secuencias génicas rrs 16S y lipL32 . Se determinó la dosis letal media en hámster inoculada por vía intraperitoneal. Se identificaron los patrones de química clínica, la duración de la leptospiremia, la leptospiruria y la histopatología, comparados con el mismo modelo inoculado con la cepa de Leptospira interrogans (Fiocruz L1-130). Resultados. Mediante análisis molecular se determinó que el aislamiento correspondía a la especie patógena Leptospira santarosai . La bacteria se recuperó a partir de tejido de riñón y de pulmón, y se detectó por medio de PCR lipL32 en el tercer día después de la infección. La proteína C reactiva aumentó en el quinto día después de la infección (3,25 mg/dl; valor normal: 0,3 mg/dl) con una disminución en el día 18 (2,60 mg/dl; valor normal: 0,8 mg/dl). Los biomarcadores de urea mostraron alteraciones indicativas de falla renal aguda (día 5 después de la infección: 49,01 mg/dl y día 18: 53,71 mg/dl). La histopatología mostró neumonía intersticial con diferentes grados de hemorragia, así como nefritis intersticial. Conclusión. Se identificó la presencia de la especie L. santarosai con capacidad patógena comparable con la cepa Fiocruz L1-130 de L. interrogans , de reconocida virulencia y tropismo pulmonar, en cuanto a los aspectos histopatológicos de tropismo a pulmón y riñón. Nunca antes se había evaluado en un modelo experimental un aislamiento de origen local bajo estos criterios biológicos.

Introduction: Is necessary to develop models for the study of leptospirosis. Objective: To genotype a Colombian strain of Leptospira isolated from a human with Weil´s syndrome and to evaluate its infection dynamics in the hamster experimental model. Materials and methods: Genotyping was performed by amplification and sequence analysis of the rrs 16S and lipL32 genes. The median lethal dose was determined in intraperitoneally inoculated hamsters. The patterns of clinical chemistry, the duration of leptospiremia, leptospiruria and pathological findings were studied and compared in the same animal model infected with L. interrogans (Fiocruz L1-130). Results: Molecular typing revealed that the isolate corresponded to the pathogenic species L. santarosai, which was recovered from hamsters´ kidneys and lungs and detected by lipL32 PCR from day 3 post-infection in these organs. There was a marked increase of C-reactive protein in animals at day 5 post-infection (3.25 mg/dl; normal value: 0.3 mg/dl) with decreases by day 18 (2.60 mg/dl: normal value: 0.8 mg/dl). Biomarkers of urea showed changes consistent with possible renal acute failure (day 5 post-infection: 49.01 mg/dl and day 18 post-infection: 53.71 mg/dl). Histopathological changes included interstitial pneumonia with varying degrees of hemorrhage and interstitial nephritis. Conclusion: The pathogenic species L. santarosai was identified in Colombia. Its pathogenicity as determined by tropism to lung and kidney was comparable to that of L. interrogans Fiocruz L1-130, well known for its virulence and pulmonar tropism. The biological aspects studied here had never before been evaluated in an autochthonous isolate.

Development of Transcriptional Fusions to Assess Leptospira interrogans Promoter Activity

Leptospirosis is a zoonotic infectious disease that affects both humans and animals. The existing genetic toolsfor Leptospira spp. have improved our understanding of the biology of this spirochete as well as the interaction ofpathogenic leptospires with the mammalian host. However, new tools are necessary to provide novel and useful information to the field. A series of promoter-probe vectors carrying a reporter gene encoding greenfluorescent protein (GFP) were constructed for use in L. biflexa. They were tested by constructing transcriptional fusions between the lipL41, Leptospiral Immunoglobulin-like A (ligA) and Sphingomielynase 2 (sph2) promoters from L. interrogansand the reporter gene. ligA and sph2 promoters were the most active, in comparison to the lipL41 promoter and the noninduced controls. The results obtained are in agreement with LigA expression from the L. interrogans Fiocruz L1-130 strain. The novel vectors facilitated the in vitro evaluation of L. interrogans promoter activity under defined growth conditions which simulate the mammalian host environment. The fluorescence and rt-PCR data obtained closely reflectedtranscriptional regulation of the promoters, thus demonstrating the suitability of these vectors for assessing promoter activity in L. biflexa.

Increased Immunogenicity to LipL32 of Leptospira interrogans when Expressed as a Fusion Protein with the Cholera Toxin B Subunit

Leptospirosis is one of the most widespread zoonosis in the world. The development of a recombinant leptospira vaccine remains a challenge. In this study, we cloned the Leptospira interrogans open reading frame (ORF) coding the external membrane protein LipL32, an immunodominant antigen found in all pathogenic leptospira, downstream of the highly immunogenic cholera toxin B subunit (CTB) ORF. Expression and assembly of the CTB-LipL32 fusion protein into oligomeric structures of pentameric size were observed in soluble fractions by Western blot analysis. The CTB-LipL32 protein demonstrated strong affinity for monosialotetrahexosylgaglioside (GM1-ganglioside) in an enzyme-linked immunosorbent assay (ELISA), suggesting that the antigenic sites for binding and proper folding of the pentameric CTB structure were conserved. Furthermore, antisera against LipL32 also recognized the CTB-LipL32 fusion protein, suggesting that LipL32 also conserved its antigenic sites, a fact confirmed by an ELISA assay showing soluble CTB-LipL32 recognition by sera from convalescent patients. In addition, soluble CTB-LipL32 generated higher specific titers in mice immunized without external adjuvant than co-administration of CTB with LipL32. The data presented here provide support for CTB-LipL32 as a promising antigen for use in the control and study of leptospirosis.

Chemokines expression during Leptospira interrogans serovar Copenhageni infection in resistant BALB/c and susceptible C3H/HeJ mice

The role of innate immune responses in protection against leptospirosis remains unclear. We examined the expression of the chemokines CCL2/JE (MCP-1), CCL3/MIP-1á (MIP-1á) and CXCL1/KC (IL-8) regarding resistance and susceptibility to leptospirosis in experimental mice models BALB/c and C3H/HeJ, respectively. A virulent strain of Leptospira interrogans serovar Copenhageni was used in this study. Twenty-five animals of each mouse strain of C3H/HeJ and BALB/c, were infected intraperitoneally with 106 cells. Five un-infected animals of each strain were kept as control. Mortality of C3H/HeJ mouse was observed while BALB/c mice were asymptomatic. The presence of leptospire DNA in tissues of infected animals was demonstrated by PCR. Chemokines were measured in serum, spleen, liver, kidney and lung of both strains of animals using immunoenzymatic assay (ELISA). Elevations in the levels of chemokines MCP-1 and IL-8 occurred in all organs and sera of C3H/HeJ and BALB/c infected mice. The levels of MIP-1á were lower when compared to MCP-1 and IL-8 in all analyzed organs, with a slight increase in liver and kidney. Our results indicate that the expression of inflammatory mediators can vary greatly, depending on the tissue and mouse strains. It is possible that the resistance to Leptospira can be partially correlated to the increase of MIP-1á observed in BALB/c mice, while an increasing and a sustained expression of MCP-1 and IL-8 in the lungs of C3H/HeJ mice can be correlated to the severity and progression of leptospirosis.

Leptospirosis / Leptospirosis

La leptospirosis es probablemente la zoonosis más extendida en el mundo. Se presenta más comúnmente en climas húmedos y cálidos. Es una fécción producida por una variedad de espiroqueta llamada Leptospira ínterrogans, de la cual se conocen más de 200 serovariedades. El contagio es infrecuente en el hombre, y se contrae a través del contacto directo o indirecto de la piel o mucosas con la orina de animales salvajes o domésticos infectados. Puede ser asintomática o presentarse como una enfermedad parecida a la influenza, a veces asociada a compromiso meníngeo. En una minoría de los casos se agregan ictericia, manifestaciones hemorrágicas e insuficiencia renal sintomática, constituyendo la enfermedad de Weil, que es la forma más grave y potencialmente fatal de la enfermedad. No son raros los cuadros clínicos incompletos o parciales. La transmisión tiene comúnmente lugar en ámbitos ocupacionales, recreacionales o domésticos. Los grupos ocupacionales más expuestos son aquellos que trabajan en contacto con suelos húmedos o con aguas estancadas, y en la esfera recreacional aquellos que se bañan, pescan o hacen deportes en aguas de poca movilidad. En el hogar tiene importancia la presencia de roedores, o el contacto estrecho con perros dentro de la casa. La mayoría de los casos se presenta en hombres, durante el verano u otoño. En su forma más común la leptospirosis adopta el aspecto clínico de un síndrome febril anictérico con múltiples síntomas inespecíficos, lo que hace difícil sospechar el diagnóstico. Sin embargo, la relevancia de la cefalea, fiebre o intensas mialgias en una persona con riesgo de exposición debe sugerir esta posibilidad. La inyección conjuntival, la presencia de meningitis aséptica, las alteraciones de sedimento urinario y la elevación frecuente de la CPK sérica pueden constituir claves clínicas valiosas. La presencia de ictericia y de insuficiencia renal clínicamente significativa definen el cuadro como enfermedad de Weil. El diagnóstico definitivo se hace por cultivo del agente causal, o más comúnmente por serología. El tratamiento antibiótico precoz y aún después de varios días de comienzo con penicilina, amoxicilina, ampicilina, tetraciclina o doxiciclina se ha demostrado como muy útil. Dado que el germen tiene una amplia sensibilidad, es posible que otros antimicrobianos también sean eficaces

Na, K-ATPase: a molecular target for Leptospira interrogans endotoxin

On the basis of our report that a glycolipoprotein fraction (GLP) extracted from Leptospira interrogans contains a potent inhibitor of renal Na,K-ATPase, we proposed that GLP-induced inhibition of Na,K-ATPase might be the primary cellular defect in the physiopathology of leptospirosis. The present study was designed to test this hypothesis by determining whether or not 1) GLP inhibits all the isoforms of Na,K-ATPase which are expressed in the tissues affected by leptospirosis, 2) Na,K-ATPase from leptospirosis-resistant species, such as the rat, is sensitive to GLP, 3) GLP inhibits Na,K-ATPase from intact cells, and 4) GLP inhibits ouabain-sensitive H,K-ATPase. The results indicate that in the rabbit, a leptospirosis-sensitive species, GLP inhibits with similar efficiency (apparent IC5O: 120-220 mug protein GLP/ml) all isoforms of Na,K-ATPase known to be expressed in target tissues for the disease. Na,K-ATPase from rat kidney displays a sensitivity to GLP similar to that of the rabbit kidney enzyme (apparent IC50: 25-80 and 50-150 mug protein GLP/ml for rat and rabbit, respectively), indicating that resistance to the disease does not result from the resistance of Na,K-ATPase to GLP. GLP also reduces ouabain-sensitive rubidium uptake in rat thick ascending limbs (pmol mm-1 min-1 ñ SEM; control: 23.8 ñ 1.8; GLP, 88 mug protein/ml: 8.2 ñ 0.9), demonstrating that it is active in intact cells. Finally, GLP had no demonstrable effect on renal H,K-ATPase activity, even on the ouabain-sensitive form, indicating that the active principle of GLP is more specific for Na,K-ATPase than ouabain itself. Although the hypothesis remains to be demonstrated in vivo, the present findings are compatible with the putative role of GLP-induced inhibition of Na,K-ATPase as an initial mechanism in the physiopathology of leptospirosis.

Aglutininas anti-leptospira em soros humanos do Estado do Piauí, com particular referência aos aspectos ocupacionais, 1994 a 1996 / Anti-leptospiral agglutinins of human sera in the state of Piauí, Northeast of Brazil, with special reference to its occupational aspects, 1994 to 1996

Foi investigada através de inquérito sorológico a provável associaçäo entre a infecçäo humana por diferentes sorotipos de Leptospira interrogans e a exposiçäo ao agente em diferentes grupos ocupacionais, entre trabalhadores urbanos e rurais do estado do Piauí e a provável correlaçäo entre prevalência e precipitaçÝo pluviométrica e as condiçöes de saneamento ambiental. Foi estudada a relaçäo entre outros fatores climáticos (temperatura e cobertura vegetal) e a infecçäo. Foram colhidas amostras de soro sangüíneo de pessoas aparentemente expostas, por motivo de atividade, confrontadas com amostras obtidas de pessoas aparentemente näo expostas (grupo controle), no período de maio de 1994 a setembro de 1996; submetidas à prova de microtécnica da soroaglutinaçäo microscópica aplicada à leptospirose. Observou-se associaçäo significante entre a prevalência e o exercício de atividades de risco mas näo entre a exposiçäo às condiçöes de saneamento ambiental e a infecçäo. As atividades de risco identificadas foram: bovinocultura (de corte e de leite), equideocultura, suinocultura, abate animal e agricultura. A maior prevalência foi encontrada na regiäo de Parnaíba (litoral) e a menor em Säo Joäo do Piauí. Dentre os sorotipos saprófitas, mais freqüente foi patoc e dentre os patogênicos, hardjo e dentre as categorias, a que apresentou risco mais elevado foi equideocultura. Observou-se correlaçäo entre prevalência e a precipitaçäo pluviométrica.

Leptospirosis: presentación de un caso clínico / Leptospirosis: a clinical case report

A propósito de un caso clínico diagnosticado en 1995 en el Hospital San Juan de Dios, se revisa el tema de las leptospirosis. A pesar de ser una afección infrecuente de nuestro país, se destacan sus características clínicas y su importancia relativa en el diagnóstico diferencial de los síndromes febriles prolongados y de las ictericias. La leptospirosis corresponde a una enfermedad infecciosa aguda que se caracteriza por una extensa vasculitis provocada por una espiroqueta del género Leptospira. Es una zoonosis de distribución mundial y que puede infectar ocasionalmente al hombre por contacto directo con el reservorio animal o por transmisión indirecta, desarrollando la enfermedad. Típicamente se considera como una afección bifásica, con una etapa inicial septicémica y una segunda fase inmune. Se describen las formas clínicas de la enfermedad, las que se pueden clasificar en dos grupos: leptospirosis anistéricas que corresponden a la forma más común y benigna (90 porciento de los casos) y leptospirosis ictéricas o síndrome de Weil (10 porciento de los casos). Se consideran aspectos diagnósticos de la enfermedad el cual se realiza principalmente por cultivo en sangre, LCR y orina y serología. Se mencionan alternativas de tratamiento. La penicilina sódica en dosis habituales es el antibiótico de elección. En general, la enfermedad es de curso benigno cuando el diagnóstico y el tratamiento son precoces. En las formas graves la mortalidad alcanza hasta el 10 porciento

La importancia de la leptospirosis humana en México: detección de anticuerpos antileptospira en una población de donadores de sangre / The importance of human leptospirosis in Mexico: detection of antileptospira antibodies in a population o blood donors

La detección de anticuerpos específicos en suero se ha utilizado como prueba diagnóstica para leptospirosis humana. En el hombre, la presencia de anticuerpos solamente se puede deber a un proceso infeccioso, ya que no existen vacunas que estimulen al sistema inmune para la producción de éstos. En el presente estudio de analizaron 206 sueros de adultos donadores de sangre, con la técnica de aglutinación microscópica contra siete serovariedades de Leptospira interrogans. El 7 por ciento de los sueros fueron positivos. La distribución de éstos por serovariedad fue la siguiente: shermani 53 por ciento, canicola 33 por ciento, pyrogenes 20 por ciento, pomona 13 por ciento e icterohaemorrhagiae 6 por ciento. El grupo etario con mayor frecuencia de seropositividad fue el de 20 a 39 años. Los resultados encontrados en individuos asintomáticos muestran que la leptospirosis en México es una zoonosis más frecuente de lo se sospecha

Leptospira interrogans sorotipo pomona em caprinos: determinaçäo do fibrinogênio, proteína total, transaminases e bilirrubinas, durante infecçäo experimental / Leptospira interrogans serovar pomona in experimental infection of goats: determination of fibrinogen, total protein, transaminases and bilirrubins

Com o objetivos de estudar os aspectos laboratoriais, através da determinaçäo do fibrinogênio plasmático, proteína total plasmática, aspartato e alanina aminostransferases séricas, e bilirrubinas séricas total, direta e indireta, utilizaram-se 20 caprinos mestiços, clinicamente sadios, de ambos os sexos, com dez meses de idade e, com peso vivo médio de oito quilogramas. Os animais foram divididos aleatoriamente em dois grupos de dez: grupo "A", controle e grupo "B", experimental. Nos animais deste último grupo foram inoculados cinco mililitros, via intraperitoneal, de cultura de Leptospira interrogans sorotipo pomona (estirpe M7/87), previamente preparada. Inicialmente, as amostras sanguíneas foram colhidas a partir do 3§ dia após inoculaçäo, em intervalos de quatro dias, entre o 3§ e 15§ dia, passando para seis dias do 16§ ao 44§ dia, e finalmente para sete dias entre o 45§ e 93§ dia. A análise estatística revelou significância a nível de 5 por cento para a bilirrubina total e direta, enquanto para as demais variáveis näo houve diferenças significativas entre os tratamentos

Leptospira ictero haemorrhagiae in rats of Trinidad, B.W.I

Leptospira ictero-haemorrhagiae was found in thirty two out of one hundred and twenty (26.6 percent) rats examined in Trinidad. Of these strains, all but two were pathogenic to guinea pigs and produced the usual symptoms followed by death in six to seven days. The incidence of infection was highest in the older and heavier rats. Cultivation on artificial media was readily obtained. (Summary)

Patologia da leptospirose: ensaio de corrrelaçäo anatomo-clínica / Pathology of leptospirosis: assay with anatomoclinical correlation

Correlacionan-se as principais manifestaçöes clínicas encontradas na forma úctero-hemorrágica da leptospirose com seus achados anatomopatológicos em autópsias e biópsias, consubstanciados em vasta experiência nas três últimas décadas na cidade do Rio de Janeiro. O mecanismo patogênico básico das lesöes é analisado como decorrente de dano celular direto pela estrutura da leptospira e de seus produtos de degradaçäo, por via de alteraçöes vasculares propiciadoras de hipoxia tecidual. O quadro clínico e laboratorial de icterícia com insuficiência renal correlaciona-se com aspectos histopatológicos de hepatite colestática centrolobular e de necrose tubular aguda com nefrite intesticial. As mialgias, presentes na quase totalidade dos casos, säo justificadas pelo quadro histológico de miosite alterativa com lesöes multifocais em aproximadamente 80 por cento dos 88 espécimes estudados em material de biópsias musculares. As manifestaçöes cardiovasculares, pulmonares, digestivas e neuropsiquiátricas derivam de miocardite, pneumonite e de lesöes associadas a fenômenos congestivos e hemorrágicos

Leptospirosis en vampiros: desmodus rotundus (geoffroy saint hilaire) / Leptospirosis in ghoul: desmodus rotundus (geoffrou saint Hilaire)

Se realizó el estudio bacteriológico y serológico en 26 especínenes de Desmodus rotundus(Geoffroy Saint Hilaire), capturados en la localidad de Shansha, Huaráz-Ancash, a fin de determinar el posible rol de reservorios de Leptospiras de la especie Leptospira interrogans. Observaciones diresctas de tejido renal y hepático al microscópio de campo oscuro, mostraron la presencia de Leptospira en 6(23.07 por ciento)especímenes. Se detectaron anticuerpos contra Leptospira mediante la reacción de aglutinación microscópica (RAM)en 5(19.23 por ciento)vampiros investigados frente a L. javanica, L. pyrogenes, L. andamana y L. grippotyphosa. Del total de cultivos realizados, se aisló una cepa de Leptospira a partir del cultivo de hígado. Por los resultados obtenidos, se evidencia el rol de reservorios de Leptospira en este grupo de mamiferos, estudiados por primera vez en Perú. Se recomienda la investigación de un número mayor de especímenes, así como en otras especies de Chirópteros, principalmente para intentar el aislamiento de Leptospira y determinar la acción de los vampiros en la transmisión de esta espiroqueta al humano y animales domésticos

Severe leptospirosis in humans

In spite of major advances in the control of infectious diseases, leptospirosis remains an important cause of morbidity and mortality in the tropics and subtropics. In rural settings, during the wetter months of the year and where there is close contact with domestic livestock, the incidence of severe disease may be particularly high. Acute renal failure is the important, life-threatening complication; its prompt recognition and treatment are the most important aspects of the successful management of severe leptospirosis (AU)

Serological and ecological study of the genus Leptospira in Jamaica

In a serological and ecological study of the genus Leptospira in Jamaica, 535 sera from three species of livestock animals, bovine, porcine and caprine were serologically tested by the microscopic agglutination technique using live leptospiral antigens. Of the number tested, 316 (59.06 percent) were reactive with titers equalling or greater than 1:100. A total of six serotypes were identified and representative serovars were as follows:- jules (serogroup Hebdomadis) 257 (51.92 percent); icterohemorrhagiae (serogroup Ictero-hemorrhagiae) 140 (28.07 percent); autumnalis (serogroup Autumnalis) 41 (8.3 percent); canicola (serogroup Canicola) 30 (6.06 percent); pomona (serogroup Pomona) 25 (5.19 percent) and abramis (serogroup Pyrogenes) 2 (0.4 percent). The porcine species recorded the highest percentage of reactive sera (65.5 percent) and the caprine the lowest (38.9 percent). The widest distribution of serotypes occurred in pigs. A comparison of the parishes showed that the positive sera from Portland (80.9 percent) and Westmoreland (71.6 percent) topped others and St. Andrew had the lowest 11.0 percent. For the ecological study, 22 soil and 25 water samples collected from five selected livestock farms in the island were cultured initially in enrichment artificial medium (Ellinghausen and McCullough semi-solid), ESS, incorporating 5-fluorouracil, for the isolation of leptospires. A total of 23 (48.9 percent) isolates were obtained:- 16 (72.7 percent) from the soil samples and 7 (28 percent) from the water samples. One water isolate was lost and the 22 parent isolates were subsequently inoculated into weanling hamsters. The kidney tissues and heartblood of the inoculated hamsters on culture yielded 8 leptospiral isolates and the microscopic agglutination test of the sera of inoculated hamsters gave 9 reactive results. There was no positive results either from culture or serological test of the control (uninoculated) hamsters. The investigations revealed that the constituents of Jamaican soil, the heavy rainfall and the various environmental factors are ideal for the survival and distribution of leptospires in the island. Flooding helps to spark off epizootic outbreaks and the leptospiral serotypes mentioned earlier are enzootic and endemic in Jamaica. The predominant serotypes are jules and icterohemorrhagiae. People at high risk are the livestock attendants and milkers. The epidemiological importance of the serotypes identified and suggested means of dealing with their adverse effects are discussed (AU)