Viable Leishmania parasites in the absence of an in vitro IFN-γ response in asymptomatic carriers.

Asymptomatic infection (the absence or inapparent signs and symptoms) has been observed in many endemic areas of leishmaniasis, however, little is known about the parasitological and immunological factors associated with this type of infection. This study aimed to identify the in vitro expression of IFN-γ in asymptomatic carriers of viable Leishmania parasites. Asymptomatic infection was identified using the Montenegro skin test in an at-risk population from Yucatan, Mexico. Parasite viability was evinced in the blood by 7SL RNA transcripts amplification. The expression of mRNA IFN-γ was analyzed in peripheral blood mononuclear cells stimulated with soluble Leishmania antigen, using RT-qPCR. Parasite viability was observed in 33.3 % (5/15) of asymptomatic subjects. No differences were found in the expression of IFN-γ between asymptomatic and healthy subjects, and no correlation was found between the presence of viable parasites and the expression of IFN-γ. This study demonstrates the persistence of Leishmania parasites in the absence of an in vitro IFN-γ response in asymptomatic carriers from Mexico.

Clinical and laboratory features of rickettsioses in Yucatan, Mexico.

Rickettsia species are obligate intracellular bacteria that can cause mild to severe human disease. Based on phylogeny, clinical symptoms, and antigenic properties, rickettsiae are classified into four groups. Infections by these agents are characterized by clinical symptoms ranging from self-limited to severe and even fatal febrile illnesses, depending on the Rickettsia spp. involved, the patient's predisposition, and timely medical care. The present study aimed to characterize rickettsial diseases in Yucatan according to clinical and laboratory features appearing in medical records corresponding to 427 samples taken between 2015 and 2018. A study was conducted over the period 2015-2018 on 427 samples. Clinical and laboratory features were documented from the patients' medical records. For molecular diagnosis, blood was collected in 3.8 % sodium citrate as anticoagulant, and DNA was extracted. Single-step and nested PCR amplification was performed using genus-specific primers for the rickettsial 17kDa and ompB genes. The amplicons obtained were purified and sequenced. A total of 22.7 % (97/427) positive cases of Rickettsia spp. were identified by PCR from 14.15 % (15/106) of the municipalities in Yucatan. 75.2 % (73/97) of the cases were from the city of Merida during the autumn (September-December). The age groups with the highest frequency of confirmed cases were pediatric (5-14 years) (57.7 %) and adults (25-49 years) (42.2 %). There were six fatal cases in children, one associated with R. typhi and five with R. rickettsii. In non-fatal cases, 32.9 % (32/97) corresponded to the spotted fever group (SFG), and 60.8 % (59/97) to the typhus group (TG). Significant differences in signs, and laboratory data, were observed between the pediatric and adult populations. For the treatment of patients, oral and intravenous doxycycline was used in severe hospitalized cases. Typhus group and spotted fever group Rickettsiae are endemic pathological agents found in urban and rural areas of our region. Molecular identification allows for greater diagnostic accuracy and timely treatment and consequently a better prognosis. It is necessary to implement or reinforce measures focused on the dissemination of knowledge regarding rickettsial diseases and their prevention.

Viable Leishmania parasites in the absence of an in vitro IFN-γ response in asymptomatic carriers

ABSTRACT Asymptomatic infection (the absence or inapparent signs and symptoms) has been observed in many endemic areas of leishmaniasis, however, little is known about the parasitological and immunological factors associated with this type of infection. This study aimed to identify the in vitro expression of IFN-γ in asymptomatic carriers of viable Leishmania parasites. Asymptomatic infection was identified using the Montenegro skin test in an at-risk population from Yucatan, Mexico. Parasite viability was evinced in the blood by 7SL RNA transcripts amplification. The expression of mRNA IFN-γ was analyzed in peripheral blood mononuclear cells stimulated with soluble Leishmania antigen, using RT-qPCR. Parasite viability was observed in 33.3 % (5/15) of asymptomatic subjects. No differences were found in the expression of IFN-γ between asymptomatic and healthy subjects, and no correlation was found between the presence of viable parasites and the expression of IFN-γ. This study demonstrates the persistence of Leishmania parasites in the absence of an in vitro IFN-γ response in asymptomatic carriers from Mexico.

First detection of Leishmania DNA in Lutzomyia longipalpis sensu lato (Diptera: Phlebotominae) in southem Mexico.

Background & objectives: Lutzomyia longipalpis sensu lato is an important vector of Leishmania infantum, the causative agent of visceral leishmaniasis (VL) in Latin America. In Mexico, this species has been recorded in endemic areas of leishmaniasis transmission, but it has never been detected as infected with Leishmania sp. This study aimed to explore the presence of Leishmania DNA in Lutzomyia longipalpis s.l. from samples collected with a human baited trap from an endemic region of leishmaniasis in southeastern Mexico. Methods: This is a prospective study where a total of 45 specimens of Lu. longipalpis s.l. collected in two sites of Yucatan state with records of leishmaniasis were tested. The nuclear ribosomal Internal Transcribed Spacer was amplified for the detection of Leishmania DNA. Results: Two females were positive for Leishmania DNA. None of the specimens positive for parasite DNA were found fed or gravid. Our finding represents the first record of infection by Leishmania in Lu. longipalpis s.l. for the country. Interpretation & conclusion: More studies are necessary to understand the potential role of this vector species in the transmission cycle of the causative agent of leishmaniasis in the southeastern and other regions of Mexico.

Leishmania (Leishmania) mexicana Infection in Wild Rodents from an Emergent Focus of Cutaneous Leishmaniasis in Yucatan, Mexico.

In 2015, emergent cases of localized cutaneous leishmaniasis (LCL) were reported in Tinum, Yucatan, Mexico. As part of an eco-epidemiological study to characterize the elements that trigger Leishmania infection in that area, we conducted a field study to investigate the occurrence of Leishmania infection in wild rodents. From November 2019 to February 2020, rodents were caught from three sites located in the municipality of Tinum, Yucatan. For each specimen, clinical signs suggestive of Leishmania infection were recorded. Samples from the tail, liver, and spleen were taken for the identification of Leishmania DNA by PCR. Twenty rodents belonging to two species were caught including Heteromys gaumeri (55%, 11/20) and Ototylomys phyllotis (45%, 9/20). Fifty-five percent of the animals presented white spots on the tail, 15% had splenomegaly, and 5% had hepatomegaly. Fifty-five percent (11/20) of the animals were found infected by Leishmania. Heteromys gaumeri was caught in all trapping sites and was the most infected species (63.6%, 7/11). The percentage of infection for O. phyllotis was 44.4% (4/9). Leishmania (Leishmania) mexicana was identified as the infecting species in two H. gaumeri. This study provides, for the first time, evidence of Leishmania infection in wild rodents from the Yucatan state. Heteromys gaumeri and O. phyllotis may be involved in the transmission cycle of L. mexicana in this emergent focus; however, further longitudinal studies are needed to confirm their role as primary reservoirs.

Urban ecology of hosts and vectors of Rickettsia in a rickettsiosis-endemic city of the Yucatan peninsula, Mexico.

Rickettsioses are vector-borne zoonotic diseases that occur in urban environments. Currently, they are associated with the presence of domestic and synanthropic animals, the ectoparasites that they harbor, and their local habitat. The implementation of prevention actions relies on the understanding of the local ecology of interactions between hosts, vector species, and the etiologic agents. In this context, this study aimed to explore and describe the occurrence of infected mammals and their ectoparasites in human urban dwellings, and those characteristics of urban dwellings associated to the presence of Rickettsia infected animals in groups of households where at least one human case of rickettsiosis has occurred in the previous year of the study. Briefly, blood-samples and ectoparasites from synanthropic and domestic animals, were obtained from groups of households from different areas of an urban settlement. Serologic and molecular diagnostics helped to identify Spotted Fever Group (SFG) and TG (Typhus Group) Rickettsia in animal and ectoparasite samples. A total of 99 mammals were sampled, 29 opossums (Didelphis virginiana), 13 house mice (Mus musculus), seven black rats (Rattus rattus) and 50 dogs. Infection occurrence in opossums was 8.3% of SFG, 50% for TG, and 4.2% of undetermined group. For house mice 46.2% for SFG and 30.8% were undetermined. Black rats 28.6% of SFG and 57.1% undetermined. Finally, dogs were 19.1% of SFG, 57.4% to TG, and 23.4% belonged to undetermined group. A total of 424 ectoparasites were collected from the mammals. In opossums occurred the ticks Ambyomma sp., Ornithodoros (Alecterobius) nr. talaje, and the flea Ctenocephalides felis. In dogs we found the ticks Rhipicephalus sanguineus s. l., Amblyomma sp., O. (A.) nr. talaje, and the flea Ct. felis. No ectoparasites were collected from rodents. The occurrence of infected animals was associated primarily with the material of the backyard floor, the type of sanitary system in the household, the presence of garbage in the backyard, presence of firewood storage, stored polyethylene terephthalate (PET) containers for sale to recyclers, and the store of construction supplies in the backyard. Nonetheless a generalized linear model showed that the household with a backyard with a dirt floor or other non-concrete material has more chances of harboring infected animals (RR= 1.74, 95% CI= 1.07-2.84 and RR= 1.03, 95% CI= 0.39-2.32 respectively). In contrast, when the house has a sanitary system of urban sewer system or a latrine outside de house, the chances of having infected animals decreased significantly (RR= 0.39, 95% CI= 0.12-0.94 and RR= 0.46, 95% CI= 0.03-2.22). We conclude that both SFG and TG rickettsioses occur in animals and their ectoparasites in peridomiciles of urban households were at least one human rickettsiosis case had occurred.

Rickettsia spp. en garrapatas (Acari: Ixodidae) que infestan perros de una comunidad rural con antecedentes de rickettsiosis, Yucatán, México / Rickettsia spp. in ticks (Acari: Ixodidae) infesting dogs from a rural community with rickettsial diseases reports, Yucatan, Mexico

RESUMEN Introducción: Las bacterias del género Rickettsia son agentes causales de las rickettsiosis, enfermedades zoonóticas transmitidas por ectoparásitos. En Yucatán, ocurrió un brote en habitantes de Bolmay, Valladolid; sin embargo, no se identificó el probable artrópodo vector. Objetivo: Analizar la presencia de Rickettsia spp. en garrapatas que infestan perros domésticos de Bolmay. Material y métodos: Se trabajaron 105 viales con hasta ocho garrapatas. Se extrajo ADN de los ectoparásitos, posterior a la identificación del estadio de desarrollo vital, sexo, género y especie. Se realizaron dos PCR para aislar fragmentos de los genes htrA (17-kDa) y rOmpB, pertenecientes a Rickettsia spp. Los productos positivos fueron secuenciados y analizados con la herramienta BLAST y el algoritmo Megablast. Resultados: Se utilizaron 291 garrapatas de los géneros Amblyomma (55.7%, 162/291), Rhipicephalus (34%, 99/291) e Ixodes (10.3%, 30/291), y las especies Rhipicephalus sanguineus sensu lato, Ixodes affinis y Amblyomma mixtum. La positividad por PCR fue de 11.4% (12/105) para los géneros Amblyomma (66.7%, 8/12), Rhipicephalus (25%, 3/12) e Ixodes (8.3%, 1/12), y las especies A. mixtum, Rh. sanguineus s. l. e I. affinis. Se identificó ADN rickettsial en ninfas, larvas, adultos, machos y hembras. Cada análisis BLAST arrojó un 96% de cobertura e identidad con varias secuencias de Rickettsia spp. como Rickettsia typhi, Rickettsia felis y Rickettsia prowazekii, por lo que no fue posible determinar la especie infectante. Conclusión: Las garrapatas que infestan perros de Bolmay, probablemente están involucrados en el ciclo de transmisión de Rickettsia spp. Se presenta la primera evidencia molecular de Rickettsia spp. en garrapatas I. affinis de Yucatán, México.

ABSTRACT Introduction: Bacteria of the Rickettsia genus are causal agents of the rickettsial diseases, zoonoses transmitted by ectoparasites. In Yucatan, in inhabitants from Bolmay an outbreak occurred; however, the probable vector was not identified. Objective: To analyze the Rickettsia spp. presence in ticks infesting domestic dogs from Bolmay. Material and methods: We used 105 vials with up to eight ticks. Total DNA from the ectoparasites was extracted, after the identification of the life stages, sex, gender, and species. Two PCR to isolate fragments of the htrA (17-kDa) and rOmpB genes belonging to Rickettsia spp., were performed. The positive products were sequenced and analyzed using the BLAST tool and the Megablast algorithm. Results: 291 ticks of the genera Amblyomma (55.7%, 162/291), Rhipicephalus (34%, 99/291), and Ixodes (10.3%, 30/291), and the species Rhipicephalus sanguineus sensu lato, Ixodes affinis, and Amblyomma mixtum, were used. The PCR positivity was 11.4% (12/105) for the genera Amblyomma (66.7%, 8/12), Rhipicephalus (25%, 3/12), and Ixodes (8.3%, 1/12), and the species A. mixtum, Rh. sanguineus s. l., and I. affinis. Rickettsial DNA in nymph, larva, adult, males, and females, were identified. Each BLAST analysis showed an identity and coverage of 96% with several Rickettsia spp. sequences, such as Rickettsia typhi, Rickettsia felis and Rickettsia prowazekii, so it was not possible to determine the infecting species. Conclusion: Ticks infesting dogs from Bolmay are probably involved in the transmission cycle of Rickettsia spp. The first molecular evidence of Rickettsia spp. in I. affinis ticks from Yucatan, Mexico, is presented.

Direct evidence of Rickettsia typhi infection in Rhipicephalus sanguineus ticks and their canine hosts.

Murine typhus is a rickettsiosis caused by Rickettsia typhi, whose transmission is carried out by rat fleas in urban settlements as classically known, but it also has been related to cat fleas in a sub-urban alternative cycle that has been suggested by recent reports. These studies remarks that in addition to rats, other animals like cats, opossums and dogs could be implied in the transmission of Rickettsia typhi as infected fleas obtained from serologically positive animals have been detected in samples from endemic areas. In Mexico, the higher number of murine typhus cases have been detected in the Yucatan peninsula, which includes a great southeastern region of Mexico that shows ecologic characteristics similar to the sub-urban alternative cycle recently described in Texas and California at the United States. To find out which are the particular ecologic characteristics of murine typhus transmission in this region, we analyzed blood and Rhipicephalus sanguineus ticks obtained from domestic dogs by molecular approaches, demonstrating that both samples were infected by Rickettsia typhi. Following this, we obtained isolates that were analyzed by genetic sequencing to corroborate this infection in 100% of the analyzed samples. This evidence suggests for the first time that ticks and dogs could be actively participating in the transmission of murine typhus, in a role that requires further studies for its precise description.

Caso fulminante de rickettsiosis (Rickettsia rickettsii) en una lactante del sureste de México / Fatal case of rickettsiosis in a toddler from southeastern Mexico

La fiebre manchada de las Montañas Rocosas es una enfermedad ocasionada por Rickettsia rickettsii, una bacteria transmitida por garrapatas infectadas, y que se caracteriza por fiebre, exantema, artralgias y mialgias, aunque, ocasionalmente, su presentación es inespecífica. Debido a que su evolución asemeja otras enfermedades exantemáticas, como dengue o chikungunya, su diagnóstico no es de primera intención, a pesar de que países como México tienen las características ecológicas y socioeconómicas propicias para su transmisión, con índices de mortalidad hasta de 30% en pacientes pediátricos. Esta elevada mortalidad se asocia a diagnósticos y terapia retrasados debido al desconocimiento médico acerca de la enfermedad, lo que propicia la aparición de formas atípicas y fulminantes de fiebre manchada de las Montañas Rocosas. El objetivo del presente trabajo es describir un caso clínico fulminante de fiebre manchada de las Montañas Rocosas para que sea considerada en el diagnóstico diferencial, lo cual impactaría directamente en los índices de mortalidad.

Rocky Mountain spotted fever is a disease caused by Rickettsia rickettsii, a bacteria transmitted by infected ticks. It is characterized by fever, exanthema, arthralgias and myalgias; but sometimes its clinical presentation is non specific. Due to its similarities with other exanthematic diseases like dengue or chikungunya, Rocky Mountain spotted fever is not a first line diagnosis, even though countries like Mexico show the ecologic and socioeconomic characteristics that favor its transmission, with a 30% mortality rate among pediatric patients. This mortality rate has been associated to a delayed diagnosis and therapy, due to a poor knowledge among physicians regarding this disease; this favors the occurrence of atypical and fulminant cases. The objective of this work is to describe a fulminant case of Rocky Mountain spotted fever, expecting that this disease could be later considered among the differential diagnosis which could directly impact its mortality rate.

[Fatal case of rickettsiosis in a toddler from southeastern Mexico]. / Caso fulminante de rickettsiosis (Rickettsia rickettsii) en una lactante del sureste de México.

Rocky Mountain spotted fever is a disease caused by Rickettsia rickettsii, a bacteria transmitted by infected ticks. It is characterized by fever, exanthema, arthralgias and myalgias; but sometimes its clinical presentation is non specific. Due to its similarities with other exanthematic diseases like dengue or chikungunya, Rocky Mountain spotted fever is not a first line diagnosis, even though countries like Mexico show the ecologic and socioeconomic characteristics that favor its transmission, with a 30% mortality rate among pediatric patients. This mortality rate has been associated to a delayed diagnosis and therapy, due to a poor knowledge among physicians regarding this disease; this favors the occurrence of atypical and fulminant cases. The objective of this work is to describe a fulminant case of Rocky Mountain spotted fever, expecting that this disease could be later considered among the differential diagnosis which could directly impact its mortality rate.

La fiebre manchada de las Montañas Rocosas es una enfermedad ocasionada por Rickettsia rickettsii, una bacteria transmitida por garrapatas infectadas, y que se caracteriza por fiebre, exantema, artralgias y mialgias, aunque, ocasionalmente, su presentación es inespecífica. Debido a que su evolución asemeja otras enfermedades exantemáticas, como dengue o chikungunya, su diagnóstico no es de primera intención, a pesar de que países como México tienen las características ecológicas y socioeconómicas propicias para su transmisión, con índices de mortalidad hasta de 30% en pacientes pediátricos. Esta elevada mortalidad se asocia a diagnósticos y terapia retrasados debido al desconocimiento médico acerca de la enfermedad, lo que propicia la aparición de formas atípicas y fulminantes de fiebre manchada de las Montañas Rocosas. El objetivo del presente trabajo es describir un caso clínico fulminante de fiebre manchada de las Montañas Rocosas para que sea considerada en el diagnóstico diferencial, lo cual impactaría directamente en los índices de mortalidad

Mecanismos de resistencia antifúngica de los azoles en Candida albicans. Una revisión / Mechanisms of antifungal resistance of azoles in Candida albicans. A review

Resumen Candida albicans es una levadura comensal capaz de causar una infección oportunista en hospederos susceptibles denominada candidiasis. El tratamiento para combatir la candidiasis puede ser tópico o sistémico según el tipo de infección, los antifúngicos más utilizados son los derivados imidazólicos (fluconazol, itraconazol, ketoconazol, miconazol etc.), sin embargo en la actualidad se observa una disminución en la efectividad de estos medicamentos, es decir, un fenómeno de resistencia de parte del microorganismo a estos fármacos, esto debido principalmente, al surgimiento de levaduras resistentes, a la aparición de nuevas especies patógenas, a la prescripción irracional de antimicóticos como profilaxis y al aumento de las dosis terapéuticas. Existen dos mecanismos por los que Candida puede adquirir resistencia a un azol. El primero es por mutaciones moleculares de la enzima diana del antifúngico, como la alteración de las enzimas relacionadas en la síntesis del ergosterol y el segundo por la alteración en las bombas de expulsión: ATP-binding cassette (ABC) y facilitadores mayores (MF). En este trabajo se resumen los principales mecanismos de resistencia en Candida y la importancia de hacer pruebas de susceptibilidad con el fin de brindar un tratamiento adecuado para este tipo de infecciones oportunistas.

Abstract Candida albicans is a commensal yeast capable of causing an opportunistic infection called candidiasis in susceptible hosts. Treatment to combat Candida may be topical or systemic according to the type of infection and the imidazole derivatives (fluconazole, itraconazole, ketoconazole, miconazole, etc.) are the antifungals most widely used. However, resistance to these drugs is observed by a decrement in their effectiveness. This is mainly due to the emergence of resistant yeasts and of new pathogenic species, as well as to the irrational prescribing of antifungal prophylaxis and the use of higher therapeutic doses. There are two mechanisms by which Candida can acquire an azole resistance, the first is by molecular mutations of antifungal target enzyme, as the alteration of enzymes related to the synthesis of ergosterols and the second by change in the efflux pumps, such as those of ATP-binding cassette and the higher facilitators. In this work the main mechanisms of resistance to Candida and the importance of performing susceptibility tests in order to provide an adequate treatment for this type of opportunistic infections are summarized.

Detection of Rickettsia felis in Wild Mammals from Three Municipalities in Yucatan, Mexico.

The aim of this study was to provide information of the occurrence of Rickettsia felis in wild mammals from three municipalities in Yucatan, Mexico. The reactivity of rodent serum to Rickettsia antigens was detected in 80.9% (17 of 21) samples using immunofluorescence assay. Polymerase chain reaction identified rickettsial DNA in spleens of 43.5% (10 of 23) rodents and 57.1% (4 of 7) opossums. The identification of the rickettsial DNA was confirmed as R. felis by restriction fragment length polymorphism and DNA sequencing. This study comprises the first report of R. felis detection in wild mammals in Yucatan.