Public Veterinary Medicine: Public Health Rabies virus variants identified in Nuevo Leon State, Mexico, from 2008 to 2015

Efrén Jaramillo-Reyna 1Laboratorio Estatal de Salud Pública, Secretaría de Salud del Estado de Nuevo León, Guadalupe, Nuevo León, México.

Search for other papers by Efrén Jaramillo-Reyna in
Current site
Google Scholar
PubMed
Close
 BSc
,
Cenia Almazán-Marín 2Unidad de Investigación Médica en Inmunología, Hospital de Pediatría, Centro Médico Nacional Siglo XXI, Instituto Mexicano del Seguro Social Ciudad de México, México.

Search for other papers by Cenia Almazán-Marín in
Current site
Google Scholar
PubMed
Close
 MS
,
Manuel E. de la O-Cavazos 1Laboratorio Estatal de Salud Pública, Secretaría de Salud del Estado de Nuevo León, Guadalupe, Nuevo León, México.

Search for other papers by Manuel E. de la O-Cavazos in
Current site
Google Scholar
PubMed
Close
 MD, Dr Med
,
Ramón Valdéz-Leal 1Laboratorio Estatal de Salud Pública, Secretaría de Salud del Estado de Nuevo León, Guadalupe, Nuevo León, México.

Search for other papers by Ramón Valdéz-Leal in
Current site
Google Scholar
PubMed
Close
 BSc
,
Alfonso H. Bañuelos-Álvarez 1Laboratorio Estatal de Salud Pública, Secretaría de Salud del Estado de Nuevo León, Guadalupe, Nuevo León, México.

Search for other papers by Alfonso H. Bañuelos-Álvarez in
Current site
Google Scholar
PubMed
Close
 MS
,
Miguel A. Zúñiga-Ramos 3Facultad de Ciencias Biológicas. Universidad Autónoma de Nuevo León, San Nicolás de los Garza, Nuevo León, México.

Search for other papers by Miguel A. Zúñiga-Ramos in
Current site
Google Scholar
PubMed
Close
 MS
,
Martín Melo-Munguía 4Instituto de Diagnóstico y Referencia Epidemiológicos, Ciudad de México, México.

Search for other papers by Martín Melo-Munguía in
Current site
Google Scholar
PubMed
Close
 BSC
,
Mauricio Gómez-Sierra 4Instituto de Diagnóstico y Referencia Epidemiológicos, Ciudad de México, México.

Search for other papers by Mauricio Gómez-Sierra in
Current site
Google Scholar
PubMed
Close
,
Albert Sandoval-Borja 4Instituto de Diagnóstico y Referencia Epidemiológicos, Ciudad de México, México.

Search for other papers by Albert Sandoval-Borja in
Current site
Google Scholar
PubMed
Close
,
Susana Chávez-López 4Instituto de Diagnóstico y Referencia Epidemiológicos, Ciudad de México, México.

Search for other papers by Susana Chávez-López in
Current site
Google Scholar
PubMed
Close
 BSc
,
José A. Díaz-Quiñonez 4Instituto de Diagnóstico y Referencia Epidemiológicos, Ciudad de México, México.

Search for other papers by José A. Díaz-Quiñonez in
Current site
Google Scholar
PubMed
Close
 DSc
, and
Nidia Aréchiga-Ceballos 4Instituto de Diagnóstico y Referencia Epidemiológicos, Ciudad de México, México.

Search for other papers by Nidia Aréchiga-Ceballos in
Current site
Google Scholar
PubMed
Close
 PhD

Abstract

OBJECTIVE

To identify rabies virus variants (RVVs) isolated from bats and terrestrial mammals in Nuevo Leon between 2008 and 2015 and Coahuila in 2006.

SAMPLE

RVVs isolated from 15 bats and terrestrial mammals in Nuevo Leon and from a cow (Bos taurus) in Coahuila, along with 46 reference rabies virus sequences.

PROCEDURES

Antigenic characterization of the 16 isolates was performed with an indirect fluorescent antibody technique. Genomic sequencing of the nucleoprotein gene in the 16 isolates was performed with a reverse transcription PCR assay. Phylogenetic reconstruction of the 62 sequences was performed by means of Bayesian inference.

RESULTS

9 isolates from bats and 1 isolate from a domestic cat that became infected as a result of contact with a Mexican free-tailed bat all clustered in the lineage associated with Lasiurus spp in the Americas or the lineage associated with Tadarida brasiliensis mexicana. An isolate from a domestic dog was identified as a variant associated with the dog-coyote lineage. The RVV isolated from a fox clustered in an Arizona fox lineage. The 3 RVVs from skunks (Mephitis macroura) were placed in a lineage with variants isolated from spotted skunks (Spilogale putorius). The RVV isolated from the cow was clustered in a lineage associated with foxes in Texas and separate from the lineage for the fox from Nuevo Leon.

CONCLUSIONS AND CLINICAL RELEVANCE

Results reinforced the need for Mexico to implement rabies surveillance and monitoring programs for bats and wild-living terrestrial carnivores.

Abstract

OBJECTIVE

To identify rabies virus variants (RVVs) isolated from bats and terrestrial mammals in Nuevo Leon between 2008 and 2015 and Coahuila in 2006.

SAMPLE

RVVs isolated from 15 bats and terrestrial mammals in Nuevo Leon and from a cow (Bos taurus) in Coahuila, along with 46 reference rabies virus sequences.

PROCEDURES

Antigenic characterization of the 16 isolates was performed with an indirect fluorescent antibody technique. Genomic sequencing of the nucleoprotein gene in the 16 isolates was performed with a reverse transcription PCR assay. Phylogenetic reconstruction of the 62 sequences was performed by means of Bayesian inference.

RESULTS

9 isolates from bats and 1 isolate from a domestic cat that became infected as a result of contact with a Mexican free-tailed bat all clustered in the lineage associated with Lasiurus spp in the Americas or the lineage associated with Tadarida brasiliensis mexicana. An isolate from a domestic dog was identified as a variant associated with the dog-coyote lineage. The RVV isolated from a fox clustered in an Arizona fox lineage. The 3 RVVs from skunks (Mephitis macroura) were placed in a lineage with variants isolated from spotted skunks (Spilogale putorius). The RVV isolated from the cow was clustered in a lineage associated with foxes in Texas and separate from the lineage for the fox from Nuevo Leon.

CONCLUSIONS AND CLINICAL RELEVANCE

Results reinforced the need for Mexico to implement rabies surveillance and monitoring programs for bats and wild-living terrestrial carnivores.

Rabies viruses are members of the genus Lyssavirus within the Rhabdoviridae family. To date, 16 species in the genus Lyssavirus have been recognized by the International Committee on Taxonomy of Viruses.1

Rabies is responsible for at least 60,000 human deaths every year.2 Virus transmission most often occurs following a bite from an infected animal or as a result of fresh saliva from an infected animal coming into contact with a break in the skin or the mucous membranes of another mammal.3 In Mexico, where dog-to-dog transmission has been controlled through vaccination, infected bats, skunks, and foxes represent the major risk of rabies transmission to humans.

Currently, > 1,116 bats species have been identified worldwide,4 and 139 bat species have been identified in Mexico,5 of which 31 have been found to be infected with rabies virus. Thirty-seven bat species have been identified in the Mexican state of Nuevo Leon, including the insectivorous Mexican free-tailed bat (Tadarida brasiliensis mexicana), which reportedly caused the first documented human death from rabies in Mexico.6,7

Genetic variants of rabies viruses can be identified antigenically with a panel of 8 monoclonal antibodies standardized by the CDC in Atlanta.8 This panel identifies 11 reaction patterns that correspond to the major RVVs circulating in Latin American and Caribbean countries.9,10 The rabies virus is a single-stranded RNA virus with an approximately 12-kilo-base genome encoding 5 viral proteins (nucleoprotein, phosphoprotein, matrix protein, glycoprotein, and an RNA-dependent polymerase),11 and it is possible to detect rabies virus through genomic sequencing of the nucleoprotein gene.7 The nucleoprotein gene is the most widely used target for rabies diagnosis,12 and sequencing it provides important data for epidemiological and evolutionary studies of all species of Lyssavirus.13

Domestic rabies, specifically rabies caused by the dog-coyote RVV, became enzootic in 1980 in coyotes (Canis latrans) along the border between Mexico and the United States. Sometime after that, the virus was translocated to various states, including Alabama and Florida, causing a local outbreak in domestic dogs in 1994.14 However, as a result of intense oral vaccination programs in the United States, the dog-coyote RVV has been considered, on the basis of results of phylogenetic analyses, to have been eliminated from the United States since 2004.15,16 Nevertheless, new cases of rabies caused by the dog-coyote RVV were reported in areas of the Mexican states of Coahuila and Tamaulipas near the US border between 2000 and 2002. In Nuevo Leon State, rabies was detected in coyotes and dogs before 1998; however, the antigenic variants and genomic sequences of these viruses were not determined.

Three bat species—T brasiliensis mexicana, Lasiurus cinereus, and Lasiurus ega—are particularly abundant in parts of Nuevo Leon State near the US border. These areas are also home to a variety of other mammals, such as gray foxes and various species of skunks and coyotes, that are potential rabies virus reservoirs and could possibly migrate to the United States.6 Recently, increased surveillance has led to the discovery of new cases of rabies involving bats and terrestrial carnivores in Nuevo Leon State. The objectives of the study reported here were to identify the RVVs involved in these cases and establish the phylogenetic relationship of these variants with variants causing historical and recent cases of rabies in border areas between Nuevo Leon and Texas and rabies in insectivorous bats in Mexico and Latin America.

Materials and Methods

A total of 62 rabies virus sequences were used in the study. This included 15 sequences from bats and terrestrial mammals in Nuevo Leon in which rabies was diagnosed between 2008 and 2015 (Figure 1), 1 sequence from a cow (Bos taurus) in Coahuila State in which rabies was diagnosed in 2006, and 46 reference sequences from GenBank.

Figure 1—
Figure 1—

Geographic distribution of 15 bats and terrestrial mammals in Nuevo Leon in which rabies was diagnosed between 2008 and 2015.

Citation: Journal of the American Veterinary Medical Association 256, 4; 10.2460/javma.256.4.438

The 15 sequences from Nuevo Leon included 9 sequences from bats (6 from Monterrey, 2 from Montemorelos, and 1 from Santiago) and 6 sequences from terrestrial mammals (3 hooded skunks [Mephitis macroura] from Mier y Noriega, 1 domestic dog from Sabinas Hidalgo, 1 domestic cat from Garcia, and 1 gray fox [Urocyon cinereoargenteus] from Salinas Victoria). Bat species were identified by experts from the Rabies Laboratory of Nuevo Leon on the basis of a taxonomy guide specific for Mexican species.6

The 46 reference sequences consisted of 20 sequences from animals in Mexico in which rabies was diagnosed between 1990 and 2007; 20 sequences from animals in the United States, including animals living near the border between the United States and Mexico, in which rabies was diagnosed between 1986 and 2007; 4 sequences from animals in Canada in which rabies was diagnosed between 1997 and 2009; 1 sequence from an animal in Brazil in which rabies was diagnosed in 2007; and 1 sequence from an animal in El Salvador in which rabies was diagnosed in 2002 (Appendix).

For the 16 animals from Nuevo Leon and Coahuila, the diagnosis of rabies was confirmed by means of fluorescent antibody testing performed at the State Public Health Laboratory of Nuevo Leon. Antigenic characterization of the isolates was performed at the Rabies Laboratory of the Instituto de Diagnostico y Referencia Epidemiologicos in Mexico City with an indirect fluorescent antibody technique that incorporated a reduced panel of 8 monoclonal antibodies (C1, C4, C9, C10, C12, C15, C18, and C19), as described.8,10

Genomic sequencing of the nucleoprotein gene in the 16 isolates was performed at the Pathogen Genomes Laboratory of the Instituto de Diagnostico y Referencia Epidemiologicos in Mexico City. Viral RNA was extracted from brain tissue with a commercial kit,a and the nucleoprotein gene was amplified with a reverse transcription PCR assay. Primers used in the PCR assay were 550Fw (5′-ATG TGY GCT AAY TGG AGY AC-3′) and 304Rvbdg (5′-ACT AGG ATT GAC RAA GAT CTT GCT CAT-3′).17 A commercial, 1-tube systemb was used. Thermocycler conditions consisted of 1 cycle at 42°C for 60 minutes; 1 cycle at 94°C for 5 minutes; 30 cycles at 94°C for 30 seconds, 55°C for 30 seconds, and 72°C for 1 minute; and 1 cycle at 72°C for 5 minutes.

Sequencing of amplification products was performed with a commercial kitc and standard analyzer.d Sequences were editede and converted to FASTA format for use in phylogenetic analysis. The 16 nucleoprotein sequences from animals in Nuevo Leon and Coahuila and the 46 reference sequences from GenBank were edited and aligned with standard software.18,f Molecular evolutionary analyses19,g were conducted to determine an adequate model of molecular evolution. Phylogenetic reconstruction was performed by means of Bayesian inference20,h that included 5 independent runs consisting of 10,000,000 generations each and a burn-in of 25%. The best-fit model was selected on the basis of the corrected Akaike information criterion, and the resulting tree was visualizedi and edited with open-source software.j

Results

Antigenic characterization of the 16 rabies virus isolates from Nuevo Leon and Coahuila revealed that isolates from 6 Mexican free-tailed bats (T brasiliensis mexicana) were all AgV9 (Table 1). The rabies virus isolated from the domestic cat, which became infected as a result of contact with a Mexican free-tailed bat, was also antigenically typed as AgV9. The 3 isolates from skunks were all AgV8, the 1 isolate from a gray fox was AgV7, the 1 isolate from a domestic dog was AgV1, and the 1 isolate from a cow was also AgV1. Two bats identified as L ega and 1 bat identified as L cinereus had atypical reaction patterns. In summary, of the 16 rabies virus isolates, 7 were AgV9, 3 were AgV8, 1 was AgV7, and 2 were AgV1; the remaining 3 isolates had atypical reaction patterns with the reduced panel of 8 monoclonal antibodies recommended by the CDC.

Table 1—

Characteristics of RVVs isolated from 15 bats and terrestrial mammals in Nuevo Leon State in which rabies was diagnosed between 2008 and 2015 and from a cow (Bos taurus) in Coahuila State in which rabies was diagnosed in 2006.

SpeciesOriginYearFragment size (bp)Sample IDGenBank Accession No.AgV
Skunk (Mephitis macroura)Mier y Noriega2008429313 Mustelid NuevoLeon 2008KY435377AgV8
Skunk (Mephitis macroura)Mier y Noriega2008417622/08skunk NLKY435376AgV8
Mexican free-tailed bat (Tadarida brasiliensis mexicana)Montemorelos20094293588 BAT Nvo Leon 2009KY435379AgV9
Mexican free-tailed bat (Tadarida brasiliensis mexicana)Monterrey20094292100/09Qpo NLKY435384AgV9
Gray fox (Urocyon cinereoargenteus)Salinas Victoria20094291769/09 Fox Nvo Leon V1KY435374AgV7
Mexican free-tailed bat (Tadarida brasiliensis mexicana)Monterrey2011429616/11qpoMontNLKY435380AgV9
Domestic catGarcía20104295389/FELINE NUEVO LEON 2010KY435378AgV9
Southern yellow bat (Lasiurus ega)Monterrey20114295426/11qpoNLKY435381Atypical
Domestic dogSabinas Hidalgo20114294195/11SabNLKY435386AgV1
Mexican free-tailed bat (Tadarida brasiliensis mexicana)Santiago20124293304 BAT NUEVO LEON 2012KY435385AgV9
Mexican free-tailed bat (Tadarida brasiliensis mexicana)Montemorelos2012429938/12QpoNLKY435382AgV9
Mexican free-tailed bat (Tadarida brasiliensis mexicana)Monterrey20124292070/12QpoNLKY435383AgV9
Hoary bat (Lasiurus cinereus)Monterrey20124292134 BAT NUEVOLEON 2012KY435388Atypical
Southern yellow bat (Lasiurus ega)Monterrey20124294873 BAT NUEVOLEON 2012KY435387Atypical
Skunk (Mephitis macroura)Mier y Noriega20154292131MxM.macrouraNL15KY435375AgV8
Cow (Bos Taurus)Coahuila2006429MN549362Coabov06MN549362AgV1

Bp = Base pair.

For the 16 rabies virus isolates from Nuevo Leon and Coahuila, there were 429 positions in the final dataset generated by genomic sequencing of the nucleoprotein gene. Molecular evolutionary analysis of all 62 sequences illustrated the relationships between the 16 isolates and the 46 reference rabies virus sequences (Figure 2). The 9 isolates from bats and the 1 isolate from a domestic cat that became infected as a result of contact with a Mexican free-tailed bat all clustered in 2 lineages: the lineage associated with Lasiurus spp in the Americas and the lineage associated with T brasiliensis mexicana. The 6 isolates from Mexican free-tailed bats and the 1 isolate from a domestic cat were all placed within the lineage of variants isolated from insectivorous bats of Canada and the United States, whereas the 1 isolate from L cinereus and the 2 isolates from L ega were clustered with RVVs from various Lasiurus spp in the Americas. All 10 of these isolates were clearly differentiated from RVVs related to the terrestrial cycle of rabies in Nuevo Leon.

Figure 2—
Figure 2—

Phylogenetic reconstruction of RVVs isolated from 15 bats and terrestrial mammals in Nuevo Leon State in which rabies was diagnosed between 2008 and 2015 and from a cow (Bos taurus) in Coahuila State in which rabies was diagnosed in 2006, along with 46 reference rabies virus sequences. Bayesian inference was used for molecular evolutionary analysis. The values on the branches represent the probabilities of the clades.

Citation: Journal of the American Veterinary Medical Association 256, 4; 10.2460/javma.256.4.438

The isolate from a domestic dog in the city of Sabinas Hidalgo, Nuevo Leon, in 2011 was identified as a variant associated with the dog-coyote lineage. This variant was located in a lineage formed by 5 other variants: 2 from Texas in 1998 and 2004, 1 from Florida in 1994, 1 from Tamaulipas in 2002, and 1 from Coahuila in 2001 (Figure 2). The RVV isolated from a fox from Salinas Victoria clustered in a lineage associated with variants from rabies cases reported in Chihuahua (1994), Sonora (2002), and Arizona (1986). The 3 RVVs from skunks were placed in a lineage with variants isolated from a spotted skunk (Spilogale putorius) in San Luis Potosi in 2002 and previously reported in an American hog-nosed skunk (Conepatus leuconotus) in 1999.21 All of these variants were placed out of the lineage for RVVs isolated from cases in Baja California, Mexico, and in Wisconsin and Canada. Finally, the RVV isolated from a cow in Coahuila was clustered in a lineage associated with foxes in Texas in 1994 and 2007 and separate from the lineage for the fox from Nuevo Leon.

Discussion

In Nuevo Leon State, the Mexican free-tailed bat is the most common bat species and the main reservoir of rabies virus, responsible for 1 human death in 1998. This was the only documented case of a human dying of rabies in Nuevo Leon. Since then, studies of the epidemiology of rabies virus in Nuevo Leon have not been reported.7 In the present study, we isolated RVVs from 3 species of insectivorous bats (T brasiliensis mexicana, L cinereus, and L ega) that are well-known reservoirs of rabies virus in the Americas. The RVVs isolated from the 6 Mexican free-tailed bats were associated with variants isolated from T brasiliensis mexicana bats in Canada and the United States. In contrast, the 3 RVVs isolated from Lasiurus spp clustered together with a widespread lineage of variants isolated in Canada, the United States, Mexico, and Brazil. As expected, the RVV isolated from an L cinereus bat was closely related to variants harbored by the bats of this species in North America, whereas the RVVs isolated from 2 L ega bats were related to variants isolated from Lasiurus xanthinus in the United States.22 This might indicate transmission between these species in Nuevo Leon State. Therefore, it is highly probable that rabies virus is circulating in the population of L xanthinus in Nuevo Leon. Another alternative is that the population of L ega bats in Nuevo Leon is sharing viruses from localities outside Nuevo Leon.

The dog-coyote RVV has not been reported in the Mexico–United States border region since 2004 and is believed to have been eliminated.15,16,23 In Northern Mexico, this antigenic variant in dogs has not been detected for 9 years; its last report was in Tamaulipas and Coahuila between 2000 and 2002,16 and it has been 16 years since the last human case related to this variant in Tamaulipas State. Our results, however, suggested movement of the dog-coyote RVV in Nuevo Leon State. It is likely that the dog-coyote variant is circulating in the Mexico–United States border region; however, the dog vaccination campaign in Mexico has been successful in avoiding infection, with just 1 case identified in 2011 in Nuevo Leon State (4195/11SabNL). Another factor that can influence the dynamics of this RVV is migration of species between the 2 countries given the similar geographic conditions. In the present study, an RVV isolated from a fox was placed in the lineages of RVVs from Arizona foxes. Moreover, RVVs isolated from 3 skunks were closely related to variants circulating in Zacatecas, San Luis Potosi, and Chihuahua. In Mexico, 2 AgVs have been described in skunks: AgV8 and AgV10. Historically, AgV8 has been isolated both from reservoir species and from spotted skunks (S putorius) in San Luis Potosi, Aguascalientes, Jalisco, and Zacatecas. Nevertheless, in the state of San Luis Potosi, this variant has been described both in hog-nosed skunks (C leuconotus) and spotted skunks (S putorius). According to studies performed by Velasco-Villa et al23 and Loza Rubio et al24 in Mexico, the main species of skunks that acts as a reservoir for rabies is S putorius. However, other species of skunks across the country may act as a reservoir of AgV8 in Mexico, which has a bat origin. To the best of our knowledge, AgV10 is limited to Baja California Sur, and according to the literature,23 the Eastern spotted skunk (Spilogale putorius lucasana) is the only species in Mexico that is a reservoir of this antigenic variant. Although AgV10 is related to the variant harbored by skunks in California, these variants are evolutionarily distinct and, as seen in the phylogenetic reconstruction performed in the present study, clustered into different lineages, indicating an evolutionarily distant event of divergence. In Canada and the United States, the skunk species most commonly associated with rabies virus is the striped skunk (Mephitis mephitis). Interestingly, AgV1 has been eliminated in dog populations through the use of vaccination campaigns, but various skunk species in Mexico harbor this antigenic variant.16

In conclusion, results of the present study reinforced the need for Mexico to implement rabies surveillance and monitoring programs for bat populations and wild-living terrestrial carnivores. Our study found circulation of RVVs in species of bats that are widely distributed in Nuevo Leon, at least some of which have a history of aggression or contact with humans. Circulation of the dog-coyote RVV, skunk RVVS, and fox RVVS must be monitored, because animals harboring these variants may represent a public health risk. Additionally, our results confirmed circulation of rabies virus in M macroura skunks, a species that has not previously been reported positive for rabies in Mexico. Future studies involving more species are needed to determine the movement of animals with rabies on both sides of the Mexico–United States border. The cases reported in the present study raise concerns about a possible reemergence of rabies in Nuevo Leon or indicate that the disease has been underestimated. Epidemiological surveillance and control measures must be implemented to protect the population. An additional measure that could help in research on rabies in the Mexico–United States border region is joint work between the countries through the North American Rabies Management Plan.

Acknowledgments

Mauricio Gómez-Sierra and Albert Sandoval-Borja were technicians in the rabies laboratory at Instituto de Diagnóstico y Referencia Epidemiológicos, Ciudad de México at the time of the study.

No third-party funding or support was received in connection with this study or the writing or publication of the manuscript. The authors declare that there were no financial or other types of conflict of interest.

The findings and conclusions in this report are those of the authors and do not necessarily represent the official opinion of the Ministry of Health in México.

The authors thank the following members of the community health staff of the Public Health State Laboratory of Nuevo León and the Instituto de Diagnóstico y Referencia Epidemiológicos in México City for technical support: Anabella Morales Rubalcaba, Alejandra Marines, Alma Lizaran Meneses, Alejandra García Muñiz, David Herrera Hernández, Israel Animas Vargas, David Martínez Solís, and Beatriz Escamilla Ríos. They also thank Sergio Lozano-Rodríguez, MD, for help in writing the manuscript.

Nidia Aréchiga-Ceballos: https://orcid.org/0000-0002-7450-3060.

ABBREVIATIONS

AgV

Antigenic variant

RVV

Rabies virus variant

Footnotes

a.

QIAamp Viral RNA Mini Kit, Qiagen, Venlo, Netherlands.

b.

Titan One Tube RT-PCR System, Roche Diagnostics GmbH, Mannheim, Germany.

c.

BigDye Terminator v3.1 Cycle Sequencing Kit, ThermoFisher Scientific, Waltham, Mass.

d.

PRISM 3130xl Genetic Analyzer, Applied Biosystems, Foster City, Calif.

e.

ChromasPro, version 1.5, Technelysium Pty Ltd, South Brisbane, Australia.

f.

Clustal W. Multiple Sequence Alignment, Kyoto University Bioinformatics Center. Available at: www.genome.jp/tools-bin/clustalw. Accessed Oct 18, 2019.

g.

JModeltest, version 2.1, Bio-Soft.net. Available at: en.bio-soft.net/tree/MODELTEST.html. Accessed Oct 18, 2019.

h.

BEAST, version 1.8, Bayesian Evolutionary Analysis Sampling Trees. Available at: beast.community. Accessed Oct 18, 2019.

i.

Rambaut A, Drummond A. FigTree. Available at: tree.bio.ed.ac.uk/software/. Accessed Oct 18, 2019.

j.

Inkscape. Available at: inkscape.org/es/. Accessed Oct 18, 2019.

References

  • 1. International Committee on Taxonomy of Viruses. Virus taxonomy: 2017. Available at: talk.ictvonline.org. Accessed Dec 4, 2018.

  • 2. World Health Organization. WHO expert consultation on rabies: second report. Geneva: World Health Organization Press, 2013.

  • 3. Acha PN, Arambulo PV III. Rabies in the tropics. In: Kuwert E, Merieux C, Koprowski H, et al, eds. Rabies in the tropics, history and current status. Berlin: Springer-Verlag, 1985;343359.

    • Search Google Scholar
    • Export Citation
  • 4. Blumenbach JF. Order Chiroptera. In: Wilson DE, Reeder DM, eds. Mammal species of the world. Baltimore: John Hopkins University Press, 2005;312529.

    • Search Google Scholar
    • Export Citation
  • 5. Medellín RA, Arita HT, Sánchez O. Identificación de los murciélagos de México, clave de campo [Identification of bats in Mexico, field code]. 2nd ed. Mexico City, Mexico: Asociación Mexicana de Mastozoología AC, 1997;78.

    • Search Google Scholar
    • Export Citation
  • 6. Jiménez G, Zúñiga-Ramos MA, Niño-Ramírez J. A. Mamíferos de Nuevo León, México [Mammals of Nuevo Leon, Mexico]. Monterrey, Mexico: Universidad Autónoma de Nuevo León, 1999;6971.

    • Search Google Scholar
    • Export Citation
  • 7. Velasco-Villa A, Orciari LA, Juarez-Islas V, et al. Molecular diversity of rabies viruses associated with bats in Mexico and other countries of the Americas. J Clin Microbiol 2006;44:16971710.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 8. Smith JS. Rabies virus epitopic variation: use in ecologic studies. Adv Virus Res 1989;36:215253.

  • 9. Diaz AM, Papo S, Rodriguez A, et al. Antigenic analysis of rabies-virus isolates from Latin America and the Caribbean. Zentralbl Veterinarmed B 1994;41:153160.

    • Search Google Scholar
    • Export Citation
  • 10. Smith JS, Orciari LA, Yager PA, et al. Epidemiologic and historical relationships among 87 rabies virus isolates as determined by limited sequence analysis. J Infect Dis 1992;166:296307.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 11. Tordo N. Characteristics and molecular biology of the rabies virus. In: Meslin FX, Kaplan MM, Koprowski H, eds. Laboratory techniques in rabies. 4th ed. Geneva: World Health Organization, 1996;2845.

    • Search Google Scholar
    • Export Citation
  • 12. Dean DJ. The fluorescent antibody test. In: Meslin FX, Kaplan MM, Koprowski H, eds. Laboratory techniques in rabies. 4th ed. Geneva: World Health Organization, 1996;8895.

    • Search Google Scholar
    • Export Citation
  • 13. Kissi B, Tordo N, Bourhy H. Genetic polymorphism in the rabies virus nucleoprotein gene. Virology 1995;209:526537.

  • 14. Clark KA, Neill SU, Smith JS, et al. Epizootic canine rabies transmitted by coyotes in south Texas. J Am Vet Med Assoc 1994;204:536540.

    • Search Google Scholar
    • Export Citation
  • 15. Sidwa TJ, Wilson PJ, Moore GM, et al. Evaluation of oral rabies vaccination programs for control of rabies epizootics in coyotes and gray foxes: 1995–2003. J Am Vet Med Assoc 2005;227:785792.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 16. Velasco-Villa A, Orciari LA, Souza V, et al. Molecular epizootiology of rabies associated with terrestrial carnivores in Mexico. Virus Res 2005;111:1327.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 17. Trimarchi CV, Smith JS. Diagnostic evaluation. In: Press A, Jackson AC, Wunner WH, eds. Rabies. San Diego: Academic Press, 2002;308344.

    • Search Google Scholar
    • Export Citation
  • 18. Kumar S, Stecher G, Tamura K. MEGA7: Molecular Evolutionary Genetics Analysis version 7.0 for bigger datasets. Mol Biol Evol 2016;33:18701874.

  • 19. Posada D. ModelTest Server: a web-based tool for the statistical selection of models of nucleotide substitution online. Nucleic Acids Res 2006;34:W700W703.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 20. Drummond AJ, Suchard MA, Xie D, et al. Bayesian phylogenetics with BEAUti and the BEAST 1.7. Mol Biol Evol 2012;29:19691973.

  • 21. Aranda M, López-De Buen L. Rabies in skunks from Mexico. J Wildl Dis 1999;35:574577.

  • 22. Kuzmin IV, Shi M, Orciari LA, et al. Molecular inferences suggest multiple host shifts of rabies viruses from bats to mesocarnivores in Arizona during 2001–2009. PLoS Pathog 2012;8:e1002786.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 23. Velasco-Villa A, Reeder SA, Orciari LA, et al. Enzootic rabies elimination from dogs and reemergence in wild terrestrial carnivores, United States. Emerg Infect Dis 2008;14:18491854.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 24. Loza-Rubio E, Nadin-Davis SA, Morales E. Molecular and biological properties of rabies viruses circulating in Mexican skunks: focus on P gene variation. Rev Mex De Cienc Pecu 2012;3:155170.

    • Search Google Scholar
    • Export Citation

Appendix

Reference rabies virus sequences used for comparison with RVVs isolated from 15 bats and terrestrial mammals in Nuevo Leon State in which rabies was diagnosed between 2008 and 2015 and from a cow (Bos taurus) in Coahuila State in which rabies was diagnosed in 2006.

MexicoUnited StatesCanadaBrazil and El Salvador
GenBank Accession No.YearStateGenBank Accession No.YearStateGenBank Accession No.YearGenBank Accession No.Year
FJ2284872006TabascoAF3948761988FloridaAF3518492009GU5528232007
FJ2284932003MichoacanGU6447262004FloridaAF3518462001FJ2284922002
FJ2284912003TamaulipasGU6447232001FloridaAF3518582001  
FJ2284882004San Luis PotosiGU6447272002TexasAF3443041997  
FJ2284851999ChihuahuaGU6447402004California    
FJ2284832001ZacatecasGU6447382004Arizona    
FJ2284842002San Luis PotosiGU6447282003Texas    
FJ2285272002TamaulipasJQ6859102002Texas    
FJ2285262001CoahuilaGU6446432005Arizona    
FJ2285311991TlaxcalaFJ2285292004Texas    
FJ2285321995PueblaFJ2285301998Texas    
FJ2285332000MichoacánFJ2285281994Florida    
FJ2285182002ChiapasFJ2285391994Arizona    
FJ2285442001Baja CaliforniaFJ2285371994Texas    
FJ2285452000Baja CaliforniaFJ2285382007Texas    
FJ2285402002SonoraAF4610451998Wisconsin    
FJ2285421994ChihuahuaFJ2285411986Arizona    
FJ2286872003SonoraFJ2285362007Texas    
AY5618052001SinaloaAF3948751995California    
AY5618062001DurangoFJ2284862007New Mexico    
  • Figure 1—

    Geographic distribution of 15 bats and terrestrial mammals in Nuevo Leon in which rabies was diagnosed between 2008 and 2015.

  • Figure 2—

    Phylogenetic reconstruction of RVVs isolated from 15 bats and terrestrial mammals in Nuevo Leon State in which rabies was diagnosed between 2008 and 2015 and from a cow (Bos taurus) in Coahuila State in which rabies was diagnosed in 2006, along with 46 reference rabies virus sequences. Bayesian inference was used for molecular evolutionary analysis. The values on the branches represent the probabilities of the clades.

  • 1. International Committee on Taxonomy of Viruses. Virus taxonomy: 2017. Available at: talk.ictvonline.org. Accessed Dec 4, 2018.

  • 2. World Health Organization. WHO expert consultation on rabies: second report. Geneva: World Health Organization Press, 2013.

  • 3. Acha PN, Arambulo PV III. Rabies in the tropics. In: Kuwert E, Merieux C, Koprowski H, et al, eds. Rabies in the tropics, history and current status. Berlin: Springer-Verlag, 1985;343359.

    • Search Google Scholar
    • Export Citation
  • 4. Blumenbach JF. Order Chiroptera. In: Wilson DE, Reeder DM, eds. Mammal species of the world. Baltimore: John Hopkins University Press, 2005;312529.

    • Search Google Scholar
    • Export Citation
  • 5. Medellín RA, Arita HT, Sánchez O. Identificación de los murciélagos de México, clave de campo [Identification of bats in Mexico, field code]. 2nd ed. Mexico City, Mexico: Asociación Mexicana de Mastozoología AC, 1997;78.

    • Search Google Scholar
    • Export Citation
  • 6. Jiménez G, Zúñiga-Ramos MA, Niño-Ramírez J. A. Mamíferos de Nuevo León, México [Mammals of Nuevo Leon, Mexico]. Monterrey, Mexico: Universidad Autónoma de Nuevo León, 1999;6971.

    • Search Google Scholar
    • Export Citation
  • 7. Velasco-Villa A, Orciari LA, Juarez-Islas V, et al. Molecular diversity of rabies viruses associated with bats in Mexico and other countries of the Americas. J Clin Microbiol 2006;44:16971710.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 8. Smith JS. Rabies virus epitopic variation: use in ecologic studies. Adv Virus Res 1989;36:215253.

  • 9. Diaz AM, Papo S, Rodriguez A, et al. Antigenic analysis of rabies-virus isolates from Latin America and the Caribbean. Zentralbl Veterinarmed B 1994;41:153160.

    • Search Google Scholar
    • Export Citation
  • 10. Smith JS, Orciari LA, Yager PA, et al. Epidemiologic and historical relationships among 87 rabies virus isolates as determined by limited sequence analysis. J Infect Dis 1992;166:296307.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 11. Tordo N. Characteristics and molecular biology of the rabies virus. In: Meslin FX, Kaplan MM, Koprowski H, eds. Laboratory techniques in rabies. 4th ed. Geneva: World Health Organization, 1996;2845.

    • Search Google Scholar
    • Export Citation
  • 12. Dean DJ. The fluorescent antibody test. In: Meslin FX, Kaplan MM, Koprowski H, eds. Laboratory techniques in rabies. 4th ed. Geneva: World Health Organization, 1996;8895.

    • Search Google Scholar
    • Export Citation
  • 13. Kissi B, Tordo N, Bourhy H. Genetic polymorphism in the rabies virus nucleoprotein gene. Virology 1995;209:526537.

  • 14. Clark KA, Neill SU, Smith JS, et al. Epizootic canine rabies transmitted by coyotes in south Texas. J Am Vet Med Assoc 1994;204:536540.

    • Search Google Scholar
    • Export Citation
  • 15. Sidwa TJ, Wilson PJ, Moore GM, et al. Evaluation of oral rabies vaccination programs for control of rabies epizootics in coyotes and gray foxes: 1995–2003. J Am Vet Med Assoc 2005;227:785792.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 16. Velasco-Villa A, Orciari LA, Souza V, et al. Molecular epizootiology of rabies associated with terrestrial carnivores in Mexico. Virus Res 2005;111:1327.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 17. Trimarchi CV, Smith JS. Diagnostic evaluation. In: Press A, Jackson AC, Wunner WH, eds. Rabies. San Diego: Academic Press, 2002;308344.

    • Search Google Scholar
    • Export Citation
  • 18. Kumar S, Stecher G, Tamura K. MEGA7: Molecular Evolutionary Genetics Analysis version 7.0 for bigger datasets. Mol Biol Evol 2016;33:18701874.

  • 19. Posada D. ModelTest Server: a web-based tool for the statistical selection of models of nucleotide substitution online. Nucleic Acids Res 2006;34:W700W703.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 20. Drummond AJ, Suchard MA, Xie D, et al. Bayesian phylogenetics with BEAUti and the BEAST 1.7. Mol Biol Evol 2012;29:19691973.

  • 21. Aranda M, López-De Buen L. Rabies in skunks from Mexico. J Wildl Dis 1999;35:574577.

  • 22. Kuzmin IV, Shi M, Orciari LA, et al. Molecular inferences suggest multiple host shifts of rabies viruses from bats to mesocarnivores in Arizona during 2001–2009. PLoS Pathog 2012;8:e1002786.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 23. Velasco-Villa A, Reeder SA, Orciari LA, et al. Enzootic rabies elimination from dogs and reemergence in wild terrestrial carnivores, United States. Emerg Infect Dis 2008;14:18491854.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 24. Loza-Rubio E, Nadin-Davis SA, Morales E. Molecular and biological properties of rabies viruses circulating in Mexican skunks: focus on P gene variation. Rev Mex De Cienc Pecu 2012;3:155170.

    • Search Google Scholar
    • Export Citation

Advertisement